Diagnosis of peripheral androgen insensitivity in a male infant excretion analysis

The hypothesis of peripheral androgen insensitivity (AIS) was examined in a boy with congenital growth hormone deficiency associated with micropenis and cryptorchidism by steroid excretion analyses compared with dihydrotestosterone (DHT) receptor analyses of foreskin biopsy homogenate. Urinary T metabolite 3 alpha,17 beta-dihydroxy-5 beta-androstane (3 beta-diol) was below the limit of detection (capillary gas chromatography) at age 1 year, but on several occasions (n = 7) normal basal values were found at age 3 years (patient: 78.9 +/- 25.4 micrograms/24 h, M +/- SD; controls, n = 15: 100.5 +/- 50.2). Normal basal- and hCG-induced excretion was noted for 3 alpha,17 beta-dihydroxy-5 alpha-androstane (3 alpha-diol) at age 1 and 3 years, respectively. Additionally, basal 3 alpha-diol excretion (n = 7) at age 3 years was 66.7 +/- 21.1 micrograms/24 h, M +/- SD; controls (n = 15) 75.8 +/- 50.4. Analysis of urinary androgens might be an alternative, noninvasive procedure for the diagnosis of peripheral AIS.     

Congenital adrenal hyperplasia: surgical considerations required to repair a 46,XX patient raised as a boy

21-hydroxylase deficiency (P450 CYP21) accounts for 90% of cases of congenital adrenal hyperplasia (CAH), which is associated with abnormally low cortisol and high production of androgen precursors and is the most common cause of ambiguous genitalia. Increased androgen causes in utero virilization of the fetus, consisting of clitoral enlargement, an urogenital sinus, and labioscrotal enlargement and fusion. This is the first case in an experience covering more than 30 years, of a 46,XX patient raised as a boy. The authors report a case of a Pakistani patient born of a consanguineous union, who came to medical attention at age 3 because of severe genital ambiguity; genetic analyses showed that the child was a compound heterozygote for CAH. The surgical management of this patient consisted of (1) staged hypospadias repairs preceded by testosterone therapy, (2) creation of a bladder graft neourethra, (3) removal of müllerian structures, (4) correction of bifid prepenile scrotum, and (5) insertion of testicular prostheses. The commitment to raise a 46,XX child as a boy is a very rare event. With a series of staged complex surgical procedures and careful steroid replacement, normal secondary sexual characteristics can be achieved in these children.     

Phenotype, hormonal profile and genotype of subjects with partial androgen insensitivity syndrome: report of a family with four adult males and one child with disorder of sexual differentiation

There is little information on the molecular basis of intrafamilial and inter-familial phenotypic heterogeneity with the same androgen receptor (AR) mutation in patients with partial androgen insensitivity syndrome. A genetic analysis was performed in a large kindred with ambiguous genitalia and the genotype–phenotype correlations were analysed. The index case was brought for sex assignment. Family history revealed four other affected members who had hypospadias and varying degrees of virilisation. All the affected males had hemizygous mutations in the third exon of the AR gene (A596T). One was also found to have a heterozygous mutation in the fourth exon of the 5 alpha reductase type 2 gene (G196S). This affected male with double mutations was better virilised compared with the other affected members with a single mutation. The degree of virilisation correlated with serum testosterone levels. Gynaecomastia was not present in any of these subjects. It is concluded that the subject with dual gene defects also had higher levels of testosterone and pubertal virilsation. Testosterone levels possibly govern the degree of pubertal virilisation in subjects with A596T gene defects. It is not clear whether the better pubertal virilsation and higher testosterone are in any way causally related to the SRD5A2 gene defect.     

Steroid receptor profile in human prostate cancer metastases as compared with primary prostatic carcinoma

The steroid receptor profile in seven prostate cancer metastases was compared with the profile in seven primary prostate cancers. The secondaries were all lymph node metastases, obtained during pelvic lymphadenectomy, preceding radical prostatectomy or irradiation. Cytosol androgen receptor content was higher in metastases, whereas the nuclear androgen receptor content was only one-fourth that in primary cancer. Cytosol progesterone as well as estrogen receptor contents were markedly lower in metastases compared with primary cancer. The steroid receptor profile differed very little between primary cancer and normal tissue. Primary prostatic carcinoma is usually obtained at early stages of the disease, whereas metastases represent a dedifferentiated, more aggressive cell population. This may explain the low amounts of progesterone, estrogen, and nuclear androgen receptor levels. The total androgen receptor content was similar in metastatic and primary disease, however, with a shift towards a cytosolic predominance in metastases. Possibly androgen receptors in metastatic disease are "deactivated."     

Effect of hydrocele on appendix testis in children

The purpose of this study was to investigate the effects of an elevated hydrostatic pressure of hydrocele on the structural integrity and steroid receptor expression pattern of the appendix testis in children. Twenty-six testicular appendages were obtained from boys （aged between 13 and 79 months, mean 40 months） who underwent surgical exploration because of hydrocele or congenital inguinal hernia. The tissue sections of testicular appendages were stained with hematoxylin-eosin. Immunohistochemistry and immunofluorescence laser microscopy were performed using monoclonal mouse anti-human receptors against androgen and estrogen receptors. Patients were divided into three groups： group A （n = 8） represented patients with groin hernia without hydrocele, who served as control group; group B （n = 7） represented patients with communicating hydrocele; and group C （n = 11） represented patients with noncommunicating hydrocele. The tissue sections of appendix testis expressed both androgen and estrogen receptors in all patients in groups A and B, and epithelial destruction was not present. The presence of androgen receptor （two of 11, P 〈 0.001） and estrogen receptor （four of 11, P = 0.006） was lower and the number of appendix testes with epithelial destruction was higher （eight of 11, P-- 0.001） in group C. We demonstrated that groin hernia and communicating hydrocele did not influence the receptor expression pattern and the anatomic structure of testicular appendages, whereas noncommunicating hydrocele caused damage as indicated by the absence of steroid receptors and destruction of the epithelial surface. A better understanding of the physiological role of testicular appendages may change the indications of surgical treatment in patients with noncommunicating hydrocele.     

Clinical and molecular characteristics in 15 patients with androgen receptor gene mutations from South China

A variety of mutations in the androgen receptor (AR) gene are linked to androgen insensitivity syndrome (AIS) or sexual development disorder. Here, we studied 15 patients with various degrees of disorders of genital hypoplasia from South China. Clinical data including basal hormone level, phenotype, karyotyping and SRY gene identification were documented. Exons with flanking intronic region of the AR gene were sequenced and analysed for mutations, and a total of eight mutations were identified in the AR gene. Of eight mutations, two novel mutations c.2518G>T (p.Asp840Tyr) and c.1186G>C (p.Gly396Arg) were predicted to be damaging by SIFT and Polyphen2 online software. Previously reported mutations: c.528C>A (p.Ser176Arg), c.1789G>A (p.Ala597Thr), c.2612C>T (p.Ala871Val), c.1752C>A (p.Phe584Leu), c.171_172insCTG (p.57_58insLeu) and c.2659A>G (p.Met887Val) were also detected in our subjects. Most of them are involved in hypospadias, penis dysplasia or other disorders of sexual development. A complete AIS case (p.Phe584Leu) with female phenotype and high serum concentrations of dihydrotestosterone (DHT) was also found. This study presented a wide range of spectrum of AIS (from partial AIS to complete AIS) caused by AR mutations in South China population. It suggests that further study with larger data set need to be performed to elucidate the differences of the phenotypes in our study.     

Comparison of finasteride (proscar®), a 5α reductase inhibitor,and various commercial plant extracts in in vitro and in vivo 5α reductase inhibition

Human prostate was used as a source of 5α reductase. Compounds were incubated with an enzyme preparation and [³H]testosterone. [³H]-dihydrotestosterone production was measured to calculate 5ã reductase activity. IC₅₀ values (ng/ml) were finasteride = 1; Permixon = 5,600; Talso = 7,000; Strogen Forte = 31,000; Prostagutt = 40,000; and Tadenan = 63,000. Bazoton and Harzol had no activity at concentrations up to 500,000 ng/ml. In castrate rats stimulated with testosterone (T) or dihydrotestosterone (DHT), finasteride, but not Permixon or Bazoton, inhibited T stimulated prostate growth, while none of the three compounds inhibited DHT stimulated growth. These results demonstrate that finasteride inhibits 5α reductase, while Permixon and Bazoton have neither anti-androgen nor 5α reductase inhibitory activity. In addition, in a 7 day human clinical trial, finasteride, but not Permixon or placebo, decreased serum DHT in men, further confirming the lack of 5α reductase inhibition by Permixon. Finasteride and the plant extracts listed above do not inhibit the binding of DHT to the rat prostatic androgen receptor (concentrations to 100 μg/ml). Based on these results, it is unlikely that these plant extracts would shrink the prostate by inhibiting androgen action or 5α reductase. © 1993 Wiley-Liss, Inc.     

Fibroblast androgen receptors in patients with genitourinary anomalies

The etiology of certain disorders of sexual differentiation is unclear. The authors have examined the hypothesis that hypospadias and other disorders compatible with a defect in androgen action, such as cryptorchidism, micropenis, chordee/penile torsion, and ectopic testis, might be explained by androgen receptor abnormalities. Therefore, 25 subjects were studied who were selected only because they had one of these developmental defects, together with a predominantly male phenotype, and no readily ascertainable explanation for the defect. Four of these subjects had mixed gonadal dysgenesis with multiple genito-urinary anomalies. They were included for comparative purposes, since there is no evidence for androgen resistance in this disorder. Patients with testicular regression syndrome (gross testosterone deficiency), impaired testosterone biosynthesis (relative testosterone deficiency), 5 alpha-reductase deficiency (altered T/DHT ratio), and a family history or endocrine profile suggestive of androgen resistance, were all excluded from evaluation. Androgen receptor content (R0) and binding affinity (Kd) were measured in 26 genital or pubic skin fibroblast strains cultured from 25 affected patients using a dispersed, whole cell assay at 22 C. There was no difference in the mean androgen receptor content (approximately 10,000 sites/cell) or binding affinity (approximately 1 nM) between the patients' fibroblasts and those from 26 fibroblast strains established from 26 normal males. Moreover, there were no differences in the nuclear uptake of [3H]dihydro-testosterone into dispersed, intact fibroblasts incubated at 37 C when 11 patient and seven normal male fibroblast strains were compared.(ABSTRACT TRUNCATED AT 250 WORDS)     

Isolated micropenis reveals partial androgen insensitivity syndrome confirmed by molecular analysis

Partial androgen insensitivity syndrome （PAIS） is the milder variant of androgen receptor （AR） defects. The subtle effects of AR mutations present in a patient with micropenis, peno-scrotal hypospadias, infertility, clitoromegaly and posterior labial fusion. We studied the association of isolated micropenis with the genetic defects resulting in androgen resistance, that is, AR gene defects and 5-α reductase type 2 （SRD5A2） deficiency. We describe two cases of isolated micropenis： one in a 14-year-old boy and the other in a 3-year-old boy who was followed until he was 10 years old. There were no findings of hypospadias, cryptorchidism or gynecomastia in either of these patients. Serum gonadotrophin and androgen levels were obtained and karyotyping was done. Human chorionic gonadotropin （hCG） stimulation testing assessed the functional capacity of the testes. DNA was extracted from peripheral leukocytes, and all exons of the SRD5A2 and AR genes were amplified by polymerase chain reaction and sequenced. In both patients, baseline testosterone （T） level was low and the values were elevated after hCG testing. The sequence of the SRD5A2 gene was normal in patient 1, and a heterozygous polymorphism, V89L, was found in patient 2. Two known mutations, P390S and A870V, were identified in patients 1 and 2, respectively. Mutations in the AR gene can be associated with isolated micropenis without other features of PAIS, such as hypospadias or gynecomastia. This underlines the importance of including AR gene analysis in the evaluation of isolated micropenis with normal plasma T to ensure proper management of the patient and appropriate genetic counseling for the family.     

Androgen receptor levels and androgen contents in the prostate lobes of intact and testosterone-treated Noble rats

Plasma testosterone (T) levels were correlated with androgen receptors, tissue content of T, and 5 alpha-dihydrotestosterone (DHT) in the three anatomically-discrete prostate lobes of intact and castrated Noble (Nb) rats bearing T-filled silastic capsules. Differences in androgen receptor content and tissue androgen levels were observed among the three prostatic lobes of intact Nb rats. Total (cytosolic and nuclear) androgen receptor levels were highest in the ventral prostate followed by the dorsolateral and anterior prostate lobes. In the ventral and anterior prostate, androgen receptors were found to be equally distributed between cytosols and nuclear extracts, whereas in the dorsolateral prostate, androgen receptors were predominantly nuclear (cytosolic: nuclear = 1.5). The ventral prostate had the highest total androgen content and DHT was the major tissue androgen in all three lobes. The ratio of tissue DHT:T varied among the lobes; the highest value was observed in the dorsolateral prostate. The higher proportions of nuclear androgen receptor, as well as the elevated tissue DHT:T found in the dorsolateral prostate compared to other lobes, suggest that differences in the androgen activation process may exist between the dorsolateral prostate and other prostatic lobes. Despite lower plasma and tissue T levels, the DHT content, weight and cytodifferentiation in all lobes of T-treated castrated rats closely approximated the situation found in intact animals. Total androgen receptor levels were, however, elevated in all prostatic lobes of T-treated, castrated rats as compared to intact controls. These increases were primarily attributed to the augmented levels of androgen receptor in the nuclear extracts of the three prostate lobes. Exposure of the prostate to a constant level of T, produced by silastic implantation, might be responsible for the higher total androgen receptor levels and enhanced nuclear androgen receptor retention found in the prostates of T-treated, castrated rats.     

Direct Effects of Medroxyprogesterone Acetate on Testes: Possible Mechanisms Examined by Testicular Perfusion

The uptake of ³H-testosterone by the nuclear androgen receptor of rat testis was studied using a perfusion system which was adapted for the simultaneous perfusion of 8 testes. Following perfusion with ³H-testosterone, the major nuclear steroid in the testis of hypophysectomized rats was testosterone rather than one of its 5α-metabolites. The accumulation of ³H-testosterone in testicular nuclei was saturable, inhibited by perfusion with excess testosterone or 4 progestins (including medroxyprogesterone acetate, MPA) which are known to bind to the androgen receptor. Saturation of testicular androgen receptors with ³H-MPA could not be demonstrated due to the high non-specific binding of this non-polar steroid. However, specific binding of ³H-MPA was demonstrated by fractionation of salt-extractable testicular nuclear receptor on sucrose gradients.
Perfused rat testes were also used to examine the direct effects of MPA on testosterone secretion. When testes were perfused with MPA, 20 μg/ml (but not 1 μg/ml) both basal and hCG induced testosterone secretion were inhibited 30 % and 60 %, respectively. In conclusion, MPA could exert a direct effect on testis via the androgen receptor as it docs in other androgen responsive tissues. Another direct effect of this progestin on the testis could be by inhibiting testosterone secretion. 7his response requires high levels of MPA in the perfusion medium suggesting that this might be a pharmacological effect of this pregestin.     

Links between genetic and environmental factors and prostate cancer risk.

BACKGROUND: Genetic polymorphisms and expression of steroid receptors may explain why some individuals are more at risk of developing prostate cancer. Some risk factors often discussed are androgen stimulation, and vitamin A and D deficiency. Long CAG-repeats in exon 1 of the androgen receptor (AR) gene on the X chromosome seem to have a protective role against androgen overstimulation. Likewise, long vitamin D receptor alleles in the poly-A tract may prevent vitamin D stimulation. METHODS: Blood samples from 59 Swedish patients with sporadic prostate cancers, 59 with hereditary prostate cancer, and 34 Japanese prostate cancer patients were compared with benign controls. Tissue specimens from 37 Swedish and 23 Japanese prostate cancer patients with matching blood samples were investigated by immunohistochemical techniques. RESULTS: The number of CAG-repeats was identical in sporadic and hereditary prostate cancer patients, but the repeats were significantly shorter than in benign controls. Benign Japanese controls were similar to Swedish controls, but Japanese prostate cancers had longer repeats than did controls. Both the vitamin D and A receptor staining was stronger in Japanese than in Swedish prostate cancer specimens. Prostate cancer occurs approximately 5 years later in Japanese compared with Swedish men. CONCLUSIONS: Varying lengths of CAG-repeats of the androgen receptor cannot fully explain racial differences in clinical prostate cancer incidence. A larger content of vitamin A and D receptors may be linked to a delayed onset of clinical prostate cancer in Japanese men.     

Androgen and glucocorticoid receptors in the pollard prostate adenocarcinoma cell lines

Both the androgen and glucocorticoid receptors of the Pollard prostate adenocarcinoma cell lines (PA-II and PA-III) have been examined and characterized. Both cell lines contain distinct binding sites for 5α-dihydrotestosterone (DHT) and dexamethasone (DM). The specific binding of the ligands to the whole cell was saturable, and Scatchard analysis showed a single class of high-affinity and low-capacity sites. (³H)-DHT was bound to a cytosolic macromolecule which sedimented at 7S after sucrose density gradient centrifugation in low salt buffer, and was completely displaced by radioinert DHT. By means of competition studies, the androgen receptor could be distinguished from that for the glucocorticoids. The DNA and protein synthesis of both tumor lines were significantly enhanced after exposure to DHT.     

Estrogen signaling and disruption of androgen metabolism in acquired androgen-independence during cadmium carcinogenesis in human prostate epithelial cells

BACKGROUND: Lethal prostate cancers often become androgen-independent due to androgen receptor (AR) overexpression. The role of cadmium in prostate tumor progression was determined. METHODS: Control and cadmium-transformed prostate epithelial cells (CTPE) were compared for steroid-induced proliferation, steroid receptor expression, and androgen metabolism. RESULTS: CTPE cells showed rapid proliferation in complete medium and sustained proliferation in steroid-reduced medium. Androgens stimulated significantly less cell proliferation and AR-related genes expression in CTPE cells. 5alpha-Dihydrotestosterone increased PSA expression more effectively in control cells. Flutamide reduced 5alpha-dihydrotestosterone-stimulated growth less effectively in CTPE cells compared to control. CTPE cells showed decreased p27 expression. Estrogen receptors were overexpressed and estradiol markedly stimulated proliferation in CTPE cells. In CTPE cells 5alpha-aromatase was markedly increased, while 5alpha-reductase was decreased. CONCLUSIONS: Cadmium-induced malignant transformation stimulates androgen independence, unrelated to AR expression or activity. Increased estrogen receptor and 5alpha-aromatase expression suggest estrogen signaling may be critical to this process.     

Is hypospadias a genetic, endocrine or environmental disease, or still an unexplained malformation?

Hypospadias is one of the most frequent genital malformations in the male newborn and results from an abnormal penile and urethral development. This process requires a correct genetic programme, time- and space-adapted cellular differentiation, complex tissue interactions, and hormonal mediation through enzymatic activities and hormonal transduction signals. Any disturbance in these regulations may induce a defect in the virilization of the external genitalia and hypospadias. This malformation thus appears to be at the crossroads of various mechanisms implicating genetic and environmental factors. The genes of penile development (HOX, FGF, Shh) and testicular determination (WT1, SRY) and those regulating the synthesis [luteinizing hormone (LH) receptor] and action of androgen (5α reductase, androgen receptor) can cause hypospadias if altered. Several chromosomal abnormalities and malformative syndromes include hypospadias, from anterior to penoscrotal forms. More recently, CXorf6 and ATF3 have been reported to be involved. Besides these genomic and hormonal factors, multiple substances found in the environment can also potentially interfere with male genital development because of their similarity to hormones. The proportion of hypospadias cases for which an aetiology is detected varies with the authors but it nevertheless remains low, especially for less severe cases. An interaction between genetic background and environment is likely.     

闭经患者雌激素与雄激素对骨密度的影响

为了解原发闭经和继发闭经患者的雌雄激素水平与骨密度的关系,收集了１８～５１岁原发闭经患者９０例,继发闭经患者２６０例,并以相同年龄正常月经１８０例为对照组。分别测定其血清雌二醇和睾酮水平,测量皮质骨和松质骨骨密度。结果：原发闭经和继发闭经患者的雌激素均低于正常对照组。２１－羟化酶缺乏患者的雌激素水平稍低但雄激素远高于其它各类闭经患者和正常对照组。各类原发闭经和继发闭经患者的皮质骨和松质骨骨密度均低于同年龄正常对照组,唯独２１－羟化酶缺乏患者骨密度不但未降低反而升高３．１％。多囊性卵巢综合征患者的骨密度降低较少,它与２１－羟化酶缺乏二者均有高雄激素的临床特征。总之,原发闭经患者骨密度低于继发闭经患者,松质骨的骨密度较皮质骨更低。结果表明雌激素和雄激素缺乏均可影响骨密度,雌、雄激素缺乏越早,骨密度越低     

Hormonal regulation of prostate-specific antigen (PSA) glycoprotein in the human prostatic adenocarcinoma cell line, LNCaP.

Prostate-specific antigen (PSA) has emerged as the most useful marker for management of patients with prostate cancer. The regulation of this glycoprotein in vivo has important clinical implications. Indirect evidence indicates that the PSA glycoprotein might be regulated by androgens, and previous studies in this laboratory have demonstrated that PSA mRNA is upregulated by androgens. The current work reports a detailed study of PSA glycoprotein expression as influenced by steroid hormones in a human prostatic adenocarcinoma cell line, LNCaP. First, we have examined the steroid binding specificity of the androgen receptor in this cell line. In comparison with wild-type rat androgen receptor in prostate, the receptor in LNCaP cells has altered affinity for a number of steroids or analogs such as progesterone (R5020), antiprogesterone (RU486), two antiandrogens (cyperoterone acetate and hydroxyflutamide), and an androgen metabolite (epitestosterone). However, its affinity for androgens (mibolerone, dihydrotestosterone, and testosterone) is not changed. The receptor does not bind to the synthetic glucocorticoids (triaminolone acetonide and dexamethasone) nor to a synthetic estrogen DES (diethylstilbestrol). The change of the steroid binding specificity of the receptor is correlated with a single mutation (A----G at nucleotide #876 relative to the initiation codon) of the steroid binding domain of the receptor. The mutation and alteration of steroid-binding specificity of the androgen receptor is also correlated with PSA glycoprotein expression affected by different ligands tested. We have demonstrated that the PSA glycoprotein is upregulated by androgens and is affected by neither epidermal growth factor nor basic fibroblast growth factor. Moreover, PSA glycoprotein could be induced by R5020, estradiol, and epitestosterone; but neither glucocorticoids nor DES had any effect on PSA induction. Interestingly, although the antiandrogen, cyperotone acetate, had the ability to induce PSA, both RU486 and hydroxyflutamide could block androgen and progesterone induction of PSA glycoprotein. Therefore, we conclude that the PSA glycoprotein expression is influenced predominantly by androgens via its receptor, and the mutation of the receptor can affect the expression of this cellular gene by the steroids other than androgens.