Irène Mowszowicz

Late-Onset Adrenal Hyperplasia in Hirsutism

We studied the incidence of late-onset adrenal hyperplasia as a cause of hirsutism, its association with the major histocompatibility complex, and its clinical expression. Twenty-four of 400 women seen because of hirsutism were found to have late-onset adrenal hyperplasia, diagnosed on the basis of a high plasma level of 17-hydroxyprogesterone, and its marked increase after ACTH stimulation. The degree of hirsutism varied widely. Plasma antigen levels were high, especially the level of androstenedione, whereas 5 alpha-reductase activity, considered to be a good index of peripheral androgen utilization, showed frequent normal or low values. The 24 patients were genotyped, along with 84 family members, and plasma hormones were measured in the family members. We found a high correlation between late-onset adrenal hyperplasia and HLA antigens B14 and Aw33. Similar biologic profiles were observed in the patients and those of their siblings who were HLA identical (n = 9), confirming that late-onset adrenal hyperplasia is linked to the histocompatibility complex. These nine siblings had no hirsutism. We therefore conclude that the role of skin sensitivity to androgens is important in determining the clinical expression of this disorder.

Clinical and Molecular Characterization of a Cohort of 161 Unrelated Women with Nonclassical Congenital Adrenal Hyperplasia Due to 21-Hydroxylase Deficiency and 330 Family Members

CONTEXT Nonclassical congenital adrenal hyperplasia (NC-CAH) due to partial 21-hydroxylase deficiency is one of the most frequent autosomal recessive diseases. OBJECTIVE The aim of this study was to determine the genotype/phenotype relationship in probands and family members. PATIENTS AND METHODS A total of 161 NC-CAH unrelated women diagnosed on late-onset symptoms, mainly hirsutism, and post-ACTH 17-hydroxyprogesterone more than 10 ng/ml, and 330 of their relatives was explored. CYP21A2 was genotyped in 124 probands. RESULTS The most frequent mutation was V281L. One severe mutation was found in 63.7% of probands, and surprisingly two severe mutations in four probands. Contrasting with the absence of clinical differences, basal testosterone, and androstenedione, basal and post-ACTH 17-hydroxyprogesterone were significantly higher in probands carrying at least one severe mutation than in those with two mild mutations (P < 0.01). Among the 330 family members, 51 were homozygotes or compound heterozygotes, and 42 were clinically asymptomatic; 242 were heterozygotes and 37 unaffected. Post-ACTH 21-deoxycortisol (21dF) was significantly higher in heterozygotes than in unaffected, however, an overlap existed. In 12 heterozygotes, post-ACTH 21dF was below 0.55 ng/ml, the cutoff value usually accepted for suggesting heterozygosity. CONCLUSIONS The study of family members underlines the variable expression of NC-CAH even within a family, suggesting that modifier factors may modulate phenotype expression. Post-ACTH 21dF cannot reliably detect heterozygous subjects. Considering the high frequency of heterozygotes in the general population, it is essential to genotype the partner(s) of the patients with one severe mutation to offer genetic counseling.

Aetiological diagnosis of male sex ambiguity: a collaborative study

Abstract. A collaborative study, supported by the Biomed2 Programme of the European Community, was initiated to optimise the aetiological diagnosis in genetic or gonadal males with intersex disorders, a total of 67 patients with external sexual ambiguity, testicular tissue and/or a XY karyotype. In patients with gonadal dysgenesis or true hermaphroditism, the incidence of vaginal development was 100%, a uterus was present in 60%; uni or bilateral cryptorchidism was seen in nearly all cases of testicular dysgenesis (99%) but in only 57% of true hermaphrodites. Mean serum levels of anti-müllerian hormone and of serum testosterone response to chorionic gonadotropin stimulation were significantly decreased in both conditions, by comparison with patients with unexplained male pseudohermaphroditism or partial androgen insensitivity (PAIS). Mutations in the androgen receptor, 90% within exons 2–8, were detected in patients with PAIS. Clinically, a vaginal pouch was present in 90%, cryptorchidism in 36%. In 52% of cases, no diagnosis could be reached, despite an exhaustive clinical and laboratory work-up, including routine sequencing of exons 2–8 of the androgen receptor. By comparison with PAIS, unexplained male pseudohermaphroditism was characterised by a lower incidence of vaginal pouch (55%) and cryptorchidism (22%) but a high incidence of prematurity/intrauterine growth retardation (30%) or mild malformations (14%). Conclusion: reaching an aetiological diagnosis in cases of male intersex is difficult because of the variability of individual cases. Hormonal tests may help to discriminate between partial androgen insensitivity and gonadal dysgenesis/true hermaphroditism but are of less use for differentiating from unexplained male pseudohermaphroditism. Sequencing of exons 2–8 of the androgen receptor after study of testosterone precursors following human chorionic gonadotrophin stimulation is recommended when gonadal dysgenesis and true hermaphroditism can be excluded.

Androgen metabolism in hirsute patients treated with cyproterone acetate

Cyproterone acetate (CPA) in association with percutaneously administered estradiol has been used for the treatment of 150 hirsute patients for periods ranging from 6 months to 3 years. A spectacular clinical improvement ensued. Plasma testosterone (T) and androstenedione (A) fell from 69.0 +/- 24 to 33.0 +/- 8 and 210 +/- 103 to 119 +/- 25 ng/dl (mean +/- SD) respectively after 3 months of treatment and remained low thereafter. In contrast, T glucuronide (TG) and 3 alpha-androstanediol (Adiol) remained high during the whole course of treatment: 37 +/- 9 and 115 +/- 43 micrograms/24 h respectively. In vitro T 5 alpha-reductase activity (5 alpha-R) in pubic skin decreased from 147 +/- 34 to 79 +/- 17 fmol/mg skin after 1 year of treatment. To elucidate the discrepancy between plasma and urinary androgens levels, T production rate (PR) and metabolic clearance rate (MCR) were measured with the constant infusion technique in 7 patients before and after 6 months of treatment. PR decreased from 988 +/- 205 to 380 +/- 140 micrograms/24 h (mean +/- SD). In contrast MCRT increased from 1275 +/- 200 to 1632 +/- 360 1/24 h; this increase in MCRT explains the striking plasma T concentration fall and the high TG and Adiol excretion relative to the decrease in PR. Antipyrine clearance rate (n = 8) increased from 36.3 +/- 5.2 to 51.5 +/- 7.4 ml/min whereas 6 beta hydroxycortisol remained unchanged. In conclusion, CPA acts through several mechanisms: (1) it lowers the androgen input to the target cells by (a) depressing T production through its antigonadotropic effect and (b) accelerating T metabolic inactivation due to a partial enzymatic inducer effect on the liver; (2) at the target cell level it competes with any remaining T for the receptor binding sites; (3) the decrease in the androgen-dependent skin 5 alpha-R is a consequence of both actions of androgen suppression and androgen receptor blockade; it reinforces the antiandrogenic effect of CPA.

Inhibition of 5 alpha-reductase activity in human skin by zinc and azelaic acid.

The effects of zinc sulphate and azelaic acid on 5α‐reductase activity in human skin were studied using an in vitro assay with 1,2[3H]‐testosterone as substrate. When added at concentrations of 3 or 9 mmol/1, zinc was a potent inhibitor of 5α‐reductase activity. At high concentrations, zinc could completely inhibit the enzyme activity. Azelaic acid was also a potent inhibitor of 5α‐reductase; inhibition was detectable at concentrations as low as 0.2 mmol/l and was complete at 3 mmol/l. An additive effect of the two inhibitors was observed. Vitamin B6 potentiated the inhibitory effect of zinc, but not of azelaic acid, suggesting that two different mechanisms are involved. When the three substances were added together at very low concentrations which had been shown to be ineffective alone, 90% inhibition of 5α‐reductase activity was obtained. If this inhibition is confirmed in vivo, zinc sulphate combined with azelaic acid could be an effective agent in the treatment of androgen related pathology of human skin.

Hormonal regulation of the androgen receptor expression in human prostatic cells in culture

The regulation of the androgen receptor (AR) expression was studied using immunocytochemical and Western blot techniques on separate cultures of epithelial cells (PNT2) and fibroblasts of human prostate. In both cell types, immunocytochemistry revealed both nuclear and cytoplasmic staining. Treatment with DHT (5 x 10(-9) M) increased both the intensity of nuclear staining and the number of cells stained. The increase, observed after DHT treatment was markedly decreased by cyproterone acetate (5 x 10(-7) M), confirming a direct action of DHT via the AR. This autoregulation of AR was confirmed by Western blot, and seems to involve transcription and protein synthesis, since it was suppressed by actinomycin D and cycloheximide. In fibroblasts, known to contain an estrogen receptor, estradiol treatment (5 x 10(-7) M) also increases the AR immunostaining. In addition, coculture studies show that epithelial cells require the presence of fibroblasts for optimal expression of the AR. These results demonstrate that prostate epithelial cells and fibroblasts have retained in culture, an hormonal sensitivity correlated with the presence of specific receptors and can serve as a model for the study of hormone action in this tissue in normal or pathological conditions.

Pharmacological and molecular evidence for the expression of the two steroid 5α-reductase isozymes in normal and hyperplastic human prostatic cells in culture

Whereas the embryological development of the human prostate is clearly dependent on steroid 5α‐reductase (5α‐R) type 2 expression, the respective expression of the two known isoforms (types 1 and 2) of 5α‐R in the adult human prostate remains unclear.

Testosterone 5α-reductase activity of skin fibroblasts increase with serial subcultures☆

Abstract Cultures of skin fibroblasts from different anatomical sites have been established and testosterone 5α-reductase assayed after the 2nd, 6th and 12th subcultures. After the 2nd passage, 5α-reduction of testosterone to both dihydrotestosterone and androstanediols correlated well with that measured in direct assays performed in total skin homogenates; this is an additional evidence that different types of skin have specific levels of testosterone 5α-reductase activity. However, there was a marked increase in 5α-reductase activity with successive subcultures (X4−5 between 2nd and 12th subcultures) in all the cell strains studied. This finding could explain why data concerning 5α-reductase in cultured skin fibroblasts are often conflicting with those in skin homogenates. It emphasizes the necessity of performing enzyme assays after the same number of passages to allow comparison from strain to strain. Different hypotheses are discussed in order to explain such variations of 5α-reductase in cultured human skin fibroblasts. Keywords: androgen metabolism in skin; 5α-reductase; cultured skin flbroblasts.

Selective venous catheterization in the evaluation of hyperandrogenism

Retrograde bilateral ovarian-adrenal vein catheterization was carried out in 16 patients with plasma testosterone levels exceeding 1.4 ng/ml (4.85 nmol/l). While pelvic ultrasonography and computerized axial tomographic scan failed to locate the androgen-producing ovarian tumors, catheterization led to a diagnosis of occult ovarian tumor in 5 patients, based on the observation of an abnormally-high and unilateral ovarian-peripheral vein testosterone gradient, which was subsequently confirmed histopathologically. In one case, unilateral elevation of the adrenal-peripheral vein testosterone gradient was found, complementing the ultrasonographic finding of an adrenal mass and confirming the diagnosis of a virilizing adrenal tumor. In the other 10 patients, gradient analysis ruled out an androgen-producing tumor, leading to the identification of nontumoral hyperandrogeny, such as a severe form of the polycystic ovary syndrome in the 6 premenopausal patients and of ovarian stromal and hilus cell hyperplasia in the 4 menopausal patients. In conclusion, appropriate indication of selective catheterization may considerably reduce the need for exploratory surgery and may help in selecting the adequate surgical approach.

Transforming growth factor β in the human prostate: Its role in stromal–epithelial interactions in non-cancerous cell culture

Stromal–epithelial interactions play a critical role in prostate development, but the precise mechanisms are still unknown. Transforming growth factor‐beta (TGFβ) could be a potential mediator of these interactions, but there is as yet no clear demonstration of its role.