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The current research aims to identify the main obstacles facing the integration of children with special needs with their peers in regular schools from the point of view of school administrations and special education teachers, and whether there are significant differences in views regarding the obstacles of integration between principals and teachers. The sample is composed of (180) respondents, half of them principals and principals of basic schools included in the integration, while the other half was one of the teachers and teachers of special education in those Schools, and the sample was selected in a strict manner and in the manner of equal samples of schools belonging to the general directorates in the governorates of Erbil, Dohuk and Sulaymaniyah. The study aims to prepare a questionnaire to identify the obstacles. The researchers prepared a closed questionnaire consisting of (73) paragraphs. The data were processed using the SPSS program. The results showed: There are obstacles in the integration from the point of view of the school administrations and the teachers of the special education. There are differences of statistical significance in the human and administrative fields from the point of view of the school administrations by the teachers of special education to the obstacles of integration. Results The researchers came up with a set of recommendations and suggestions.

Item does not contain fulltext Over the past 5 yr several inactivating mutations in the LH receptor gene have been demonstrated to cause Leydig cell hypoplasia, a rare autosomal recessive form of male pseudohermaphroditism. Here, we report the identification of two new LH receptor mutations in a compound heterozygous case of complete Leydig hypoplasia and determine the cause of the signaling deficiency at a molecular level. On the paternal allele of the patient we identified in codon 343 a T to A transversion that changes a conserved cysteine in the hinge region of the receptor to serine (C343S); on the maternal allele a T to C transition causes another conserved cysteine at codon 543 in trans-membrane segment 5 to be altered to arginine (C543R). Both of these mutant receptors are completely devoid of hormone-induced cAMP reporter gene activation. Using Western blotting of expressed LH receptor protein with a hemagglutinin tag, we further show that despite complete absence of total and cell surface hormone binding, protein levels of both mutant LH receptors are only moderately affected. The expression and study of enhanced green fluorescent protein-tagged receptors confirmed this view and further indicated that initial translocation to the endoplasmic reticulum of these mutant receptors is normal. After that, however, translocation is halted or misrouted, and as a result, neither mutant ever reaches the cell surface, and they cannot bind hormone. This lack of processing is also indicated by reduced presence of an 80-kDa protein, the only N-linked glycosylated protein in the LH receptor protein profile. Thus, complete lack of signaling by the identified mutant LH receptors is caused by insufficient processing from the endoplasmic reticulum to the cell surface and results in complete Leydig cell hypoplasia in this patient.

Androgens play a crucial role in the development, maintenance and regulation of male phenotype and reproductive physiology through the androgen receptor, a transcription factor. Testosterone or dihydrotestosterone binding induces a trans-conformation of the androgen receptor and allows its translocation into the nucleus, where it recognizes specific DNA sequences. Recent developments in molecular genetics, as well as structural analysis of the androgen receptor, allow a better understanding of the structure/function relationship of this nuclear receptor. Molecular analyses of androgen insensitivity syndrome, as well as hormone-resistant prostate cancer, Kennedy's disease and isolated male infertility, have been proved useful as privileged models for this purpose. In the absence of identified AR receptor mutations in androgen insensitivity syndromes, abnormalities of transcriptional cofactor should be considered. Finally, identification of androgen-dependent genes will be helpful for evaluating the degree of the molecular defect of androgen action within target cells.

The aim of our study was to measure the volume of each carpal bone during childhood and adolescence by image processing from computed tomography (CT) scans, and to analyze the relationship between the eight carpal bones. Thirteen CT scans were performed in nine normal prepubertal, peripubertal and post-pubertal children, six boys and three girls, aged 5-14 years. Each scan was processed in order to extract the carpal bones. The volume was computed for each bone. There was a significant correlation between carpal bone volume and age (0.55 < r < 0.79), and a very strong correlation between the volume of a given carpal bone and the volume of all the others, whatever the age (0.87 < r < 0.99, p < 0.01). Image processing is a potentially useful method for assessing bone maturation. The constant ratio between carpal bone volumes indicates that these bones interact with each other in wrist bone maturation.

We previously described an androgen receptor (AR) point mutation located in the DNA-binding domain (DBD), adjacent to another AR substitution. Both were observed in two unrelated families with male breast cancer (MBC) and partial androgen insensitivity syndrome. This work was designed to determine the potential role of these two residues by in vitro study of the consequences of these two substitutions on biological functions and their structural impact at the atomic level. Mutant ARs revealed normal androgen-binding affinities and weaker DNA binding to an isolated androgen-responsive element. In cotransfection assays the mutant ARs displayed a reduced transactivation efficiency at 0.3 x 10(-10) M. Neither binding to an estrogen-responsive element nor transactivation efficiency of an ERE reporter gene was observed. Molecular modeling revealed that Arg607 and Arg608 were partially surface-exposed and located in adjacent areas in the AR-DBD complex with DNA. This is in favor of a protein-protein interaction. It is conceivable that such an interaction could be affected by mutation of one of these two arginines.

Le syndrome de Klinefelter se caracterise, chez l'adulte, par un ensemble de manifestations cliniques et biologiques typiques. Chez l'enfant, en depit d'une incidence elevee (1 a 2/1000 garcons nouveau-nes), le syndrome de Klinefelter reste peu diagnostique, de par une symptomatologie peu evocatrice: dysmorphie faciale, anomalie des organes genitaux, troubles du langage, avance staturale. Pendant ou apres la puberte, ce syndrome est reconnu devant une atrophie testiculaire, constante, une grande taille, une gynecomastie principalement. Dans la mesure ou une androgenotherapie en periode pubertaire est susceptible d'ameliorer significativement le pronostic pejoratif de ces adolescents, il faut souligner la necessite de realiser un diagnostic precoce et une prise en charge en periode pubertaire.

La determination testiculaire, premiere etape de la differenciation sexuelle masculine, est sous le controle de facteurs genetiques. Le role cle du gene SRY dans ces mecanismes est maintenant bien etabli. Cependant de nombreux arguments, cliniques et genetiques, plaident en faveur de l’intervention d’autres genes en aval ou en amont de SRY. Ainsi, la plupart des hommes 46, XX ont herite, par translocation, du gene SRY, responsable de la differenciation des gonades dans le sens testiculaire, mais dans 10% des cas le gene SRY n’est pas retrouve. De meme, si l’analyse de femmes 46, XY a participe a l’identification du gene SRY, 80% des cas de dysgenesies gonadiques ne sont pas expliques par une anomalie de SRY. Un certain nombre de situations cliniques permettent de suspecter le role des autosomes (chromosomes 1, 9, 10 17 …) et du chromosome X dans la pathologie de la determination sexuelle. Les travaux les plus avances concernent le facteur TDF-X, qui agirait comme un represseur de la determination testiculaire. Au total, les mecanismes moleculaires de la determination sexuelle apparaissent extremement complexes, faisant intervenir probablement plusieurs genes en une cascades de reactions.

Resume Les auteurs mettent en evidence l'importance et l'urgence d'un diagnostic precis de toute anomalie de la sphere genitale. Cette prise en charge passe par la collaboration de nombreux specialistes au sein d'une equipe pluridisciplinaire.

In patients with androgen insensitivity syndrome (AIS), RFLP study of the androgen receptor gene made it possible to analyze whether deletions or mutations could be responsible for abnormalities in androgen responsiveness. We studied RFLPs of DNA from 25 46,XY patients with partial AIS (PAIS), defined as a concentration of androgen receptor in genital-skin fibroblasts less than 340 fmol/mg DNA, and DNA from 27 46,XY patients with complete AIS (CAIS) with no detectable androgen receptor site. DNA samples were digested with BamHI, EcoRI, HindIII and TaqI restriction enzymes and hybridized with three cDNA probes covering the three domains of the androgen receptor. When we had the maternal and an unaffected brother's DNA, we analyzed the two androgen receptor gene polymorphisms described, the HindIII and the exon 1 CAG repeat polymorphisms, in order to distinguish the two maternal X chromosomes, and to detect carriers of AIS. We did not find any large deletion among the 52 patients. We observed a heterozygous mother in 3 of 14 families studied with the HindIII polymorphism, and in 12 of 25 families using the exon 1 CAG repeat polymorphism. This study suggests that in AIS, abnormalities in androgen receptor response could be related to point mutations or microdeletions rather than to gross structural alterations of the androgen receptor gene. Furthermore, unless the point mutation has been described, exon 1 and HindIII polymorphism studies would enable the identification of carriers in 50% of families, and the prenatal diagnosis of AIS.

The androgen receptor (AR), which mediates androgen action in the cell, belongs to the superfamily of nuclear receptors, a large group of transcription factors. Recent studies have described how the AR acts on specific target genes. The receptor's specificity of action depends on its regulation at different levels: expression in the cells, ligand binding and DNA-specific sequence recognition by structurally conserved domains and regulation by transcriptional factors in an integrated response. We propose, here, an overview of recent works on the molecular regulation of androgen-dependent genes by AR.

Androgens play a crucial role in the development, maintenance and regulation of male phenotype and reproductive physiology through the androgen receptor, a transcription factor. Testosterone or dihydrotestosterone binding induces a trans-conformation of the androgen receptor and allows its translocation into the nucleus, where it recognizes specific DNA sequences. Recent developments in molecular genetics, as well as structural analysis of the androgen receptor, allow a better understanding of the structure/function relationship of this nuclear receptor. Molecular analyses of androgen insensitivity syndrome, as well as hormone-resistant prostate cancer, Kennedy's disease and isolated male infertility, have been proved useful as privileged models for this purpose. In the absence of identified AR receptor mutations in androgen insensitivity syndromes, abnormalities of transcriptional cofactor should be considered. Finally, identification of androgen-dependent genes will be helpful for evaluating the degree of the molecular defect of androgen action within target cells.

Bone maturation is the only reliable indicator of growth and its radiologic assessment with or without automated systems is a qualitative method. Image processing allows the study of bone maturation with quantitative data. Carpal bone maturation was studied in 20 children (13 boys and 7 girls, ages ranging from 4 to 15 years) without any clinical evidence of endocrine disease by image analysis from computed tomography (CT) scans. Each wrist CT scan was processed in order to extract the carpal bones and to measure quantitative data regarding volume, axes of inertia and density for each bone. The volumes and the length of the inertia axes were significantly correlated with age. Whatever the age, there were strong correlations between the volume or the length of the main inertia axis of one carpal bone and that of all others. The decrease in the carpal bone volume measured from the processing procedure compared with the theoretical volume of bone defined from the length of the three inertia axes indicated a change in bone shape during growth. Although the mean density was constant, there was an increase in the standard deviation of density with age. Skeletal maturity assessment with image analysis from CT scans seems to be a good complementary investigation to determine bone age in children.

L'évaluation radiographique de la maturation osseuse assistée ou non par ordinateur repose sur la comparaison entre l'aspect radiographique d'un segment de membre d'un enfant avec l'aspect standardisé dans une population comparable d'enfants à différents stades de maturation. L'évaluation de la maturation osseuse par analyse d'image à partir d'examens tomodensitométriques du poignet est une méthode quantitative permettant de mesurer séparément pour chaque os du carpe le volume, les axes d'inertie et la densité moyenne. Les modifications morphologiques des os du carpe pendant la croissance sont caractérisées par des données quantitatives à partir de la longueur des axes d'inertie et des volumes. L'analyse d'image est une méthode d'avenir pour l'évaluation de la maturation osseuse pendant la croissance. © 2000 Éditions scientifiques et médicales Elsevier SAS

Hyperandrogenism in adolescent girls can be a troubling problem because of the difficulty in establishing a diagnosis and in prescribing appropriate therapy. Androgen excess in adolescent patients encompasses a spectrum of clinical presentations, including acne, hirsutism, oligomenorrhea, amenorrhea, virilism, and ovarian cysts. Androgen excess is a clinical and chemical feature of idiopathic hirsutism, late-onset forms of congenital adrenal hyperplasia, and polycystic ovarian disease; in some cases, functional hyperandrogenism is discussed. We recommend screening for hyperandrogenism by measuring blood levels of testosterone, dehydroepiandrosterone sulfate, and delta 4-androstenedione, while others propose a first dexamethasone suppression test for evaluation of free testosterone, dehydroepiandrosterone sulfate, and cortisol. Treatment will be chosen according to particular symptoms such as acne, hirsutism, obesity, or oligomenorrhea.

Molecular Analysis of the Androgen Receptor Gene in Patients with Complete and Partial Androgen Insensibility Syndrome

Top of pageAbstract Introduction. Genital abnormalities such as micropenis, hypospadias, and cryptorchidism have been reported in Klinefelter's syndrome. We studied the biochemical and molecular characteristics of the androgen receptor (AR) in 5 patients with Klinefelter's syndrome (47,XXY) and a severe micropenis. Patients. Clinical, biochemical and molecular data are reported in the table below. Methods. AR binding capacity was studied on genital skin fibroblasis and SSCP analyses were performed in exons 4-8 in order o delect any alterations within the androgen binding domain of the AR gene. Results. The 5 patients exhibited a decreased amount of AR (mean = 258±36 fmol/mg DNA vs 650±200 fmol/mg DNA for N) compatible with the diagnosis of PAIS while the Kd of the AR were in normal range (mean = 0.7±0.2 nM vs 0.6±0.3 nM for N). Furthermore, no band shifts, characteristic of point mutations, were found by PCR coupled with SSCP. Known AR mutated exons detecled by SSCP were used as control. Discussion. AR gene mutations have been reported in patients with partial androgen insensitivity syndrome and diminished receptor binding capacity. These mutations have been located within the androgen-binding domain. In these patients with Klinefelter's syndrome and severe micropenis, the decrease of AR binding capacity is in favor of PAIS. However, a diminution of AR gene expression responsible for the low amount of AR cannot be ruled out.

Introduction. Most XX males, SRY positive, have a normal male phenotype. I some cases, XX males, SRY negative, are associated with ambiguous genitalia (AG). The aim of this study was to find out whether, in these patients, AR amount is diminished in target cells and whether partial androgen resistance is relevant to molecular abnormalities of the AR gene. Patients and Method. Three XX males with AG were studied. PCR of the SRY gene, known to be equated to the testis determining gene, was negative. AR binding capacity (Bmax) and dissociation constant (Kd) of the A-AR complex were determined on genital skin fibroblasts by Scatchard analysis. PCR coupled with SSCP analysis was performed on all exons (4-8) encoding for the androgen-binding domain (ABD) of AR. Results. Receptor-binding capacity was respectively 243, 206 and 162 fmol/mg DNA (N = 650 ± 200 fmol/mg DNA). Kd was respectively 1, 0.66 and 0.4 nM (N = 0.6 ± 0.3 nM). No band shifts were detected on the 5 exons studied. Known mutated exons of AR detected by SSCP were used as control. Discussion. In patients with partial androgen insensitivity syndrome and diminished receptor binding capacity, a mutation has been reported within the ABD. In the 3 XX males, AG seems to be due to a low level of AR, without any detected mutation in the ABD. In these XX males, SRY negative, AG is relevant to a low level of expression of the AR gene. We suggest that the product of the SRY gene could thus be a transcriptional factor of the AR gene. Production of AR by AR-negative COS cells transfected by SRY gene is now under investigation.

Introduction. AR belongs to a family of ligand-induced transcription factors, and its normal function is to control the differentiation, development and maintenance of male reproductive function. Complete (CAIS) and partial (PAIS) androgen insensitivity syndrome are X-linked disorders occuring in 46,XY patients and leading respectively to a female phenotype and an undermasculinized male. We have determined the AR binding capacity on genital skin fibroblasts and studied the AR gene in the DNA of 5 patients with CAIS and one patients with PAIS. Families. All patients with CAIS were characterized by a referred to our clinic for genital bud, cryptorchidism and perineal hypospadias. Two maternal uncles had ambiguous genitalia. Methods. AR binding capacity was determined on genital skin fibroblasts (GSF) using [3H]-R 1881. Point mutations were detected by SSCP analysis of exons 2-7 of the AR gene on DNA from white blood cells and identified after direct DNA sequencing. Results. AR binding capacity (Bmax). dissociation constant (Kd) and detected mutations are reported in the table below. Normal Bmax = 65 ± 200 fmol/mg DNA. Normal Kd = 0.6 ± 0.3 nM. Discussion. In these families, we detected only amino acid substitution. In two cases (indicated by †), a new restriction site was created which made possible the carrier diagnosis of patients's sisters. Patients 3 presented a de novo mutation. In 2 cases (indicated by 0) a restriction site was abolished and carrier diagnosis was possible. In family 6, we found a mutation reported threefold indicating a hot-spot amino acid. As expected, the mutation described within the DNA-binding domain did not alter the AR binding capacity (patients 1 and 2). Further receptor studies are currently underway to support the potential androgen-AR and DNA-AR affinity changes.

Understanding Turner syndrome spontaneous adult height is a prerequisite to an accurate assessment of the therapeutic efficiency of growth hormone treatment. The heights described in the literature reveal significant differences (136 to 147 cm). Our collaborative study pooled results from 16 pediatric endocrinology centers and obtained a great number of spontaneous adult heights (n = 216). The selective criteria were: chronological age (CA)>18 years, bone age (BA)>16 years, typical kariotype, no treatment with growth hormone nor anabolic steroids. Mean CA was 23.3 ± 5.6 years. Chromosomal anomalies were: monosomy X: 56.7%, mosaicism: 32.7%, structural aberration: 10.6%. Mean adult height in the whole group was: 141.5 ± 6.6 cm). There was no significant difference as to monosomy X (141.1 ± 6.4 cm for n = 121); mosaicism (141.5 ± 7.5 cm torn = 72); anomaly X (141.4 ± 5.0 cm for n = 23). Mean parental height was 170.4 ± 7.1 cm (father) and 160.1 ± 6.2cm (mother). Parental height and patients heights correlated significantly: more so with fathers' heights (r = 0.50) than with mothers' (r = 0.42). The correlation was still clearer with the target height (r = 0.55). Individual patients heights varied greatly (ranged irom 129 to 161 cm). The analysis heights in the 2 groups: > 146.6 cm (+ 1 SDS) n = 34 and < 134.8cm (-1 SDS) n = 35 showed no difference in chromosomal anomaly distribution but revealed a very significant difference in parental height (158.3 ± 3.6 cm and 170.4 ± 4.9 cm respectively). These results showed: Turner syndrome adult heights were 4 SD below mean French heights, no difference in kariotype, a strong correlation with parental height and target height. Individual height differences can be explained by parental heights.

ABSENCE OF MUTATION IN THE G PROTEIN a s SUBUNIT GENE IN PATIENTS WITH Mc CUNE ALBRIGHT SYNDROME (MAS)

Androgen dependent skin disorders are important in clinical practice. Effective topical antiandrogens would lead to a breakthrough in their treatment. Although many attempts have been performed to develop such compounds, major successes have not been forthcoming. In the present study three existing antiandrogenic molecules have been compared with regard to their effect on androgen metabolism, receptor competition and on histological parameters in the hamster flank organ test. It appears that the effect on the hamster pigmented spot can be predicted on the basis of molecular mechanism. However, the effects on histological parameters are apparently dependent on additional factors such as metabolism of the active substance before reaching the sebaceous structure or limited penetration through the skin surface. The results indicate that in the development of new antiandrogens pre-screening can be performed with the aid of metabolic and receptor studies, while the histological parameters in the hamster flank organ test provide an animal model with a good predictive value.

Estrone and estradiol (E1 + E2) concentrations in saliva were compared with four other parameters of estrogen status in five normal ovulatory women and ten FSH stimulated women selected for an in vitro fertilization program. E1 + E2 in saliva, plasma, and urine were assessed by a rapid, specific and sensitive enzymatic assay using bioluminescence. The free fraction of plasma estradiol was determined by equilibrium dialysis and total plasma estradiol by conventional radioimmunoassay. The pattern of E1 + E2 variation in saliva was similar to that of free plasma estradiol and the two parameters were correlated in both spontaneous and stimulated cycles. However, the lower correlation coefficient (r = 0.52, P less than 0.001) in spontaneous cycles compared with the high (r = 0.96, P less than 0.001) in the stimulated cycles shows that salivary E1 + E2 could be representative of plasma free estradiol in stimulated cycles but not in normal cycles. The free fraction of plasma estradiol reproduced the variation of total plasma estradiol in spontaneous as well as in FSH stimulated cycles and both parameters were strongly correlated (r = 0.91, P less than 0.001 and r = 0.90, P less than 0.001), respectively. The data show that salivary E1 + E2 concentrations are highly representative of the free fraction of E2 in plasma and at a lesser extend (r = 0.72, P less than 0.001) of total plasma E2 in FSH stimulated cycles.

The benign hyperplasia of the prostate is a manifestation of aging, involving the accumulation, within the gland, of dihydrotestosterone, the probable mediator of the hyperplasia. Binding studies were performed on the cytosolic androgenic receptor of the rat prostate using [3H]methyltrienolone as a ligand. The binding of [3H]methyltrienolone at 5 nM, was inhibited by various drugs, such as methyltrienolone and cyproterone acetate. Permixon, a liposterolic extract of the plant, Serenoa Repens B, inhibits competitively the binding to the cytosolic receptor of the rat prostate. Various vegetable and mineral oils, the plant steroid: beta sitosterol and the antiprostatic drug: Tadenan, were all found to be inactive. The antiprostatic activity of Permixon shown in animal studies and controlled clinical trials, may thus result from a direct action at the cytosolic receptor.

The effect of human somatomedin C/insulin-like growth factor I(SM-C/IGF-1) and human growth hormone (hGH) on colony formation by erythroid precursor cells (CFU-E and BFU-E) from children's bone marrow or blood was studied by methylcellulose cloning assay. We found that physiological concentrations of IGF-1, but not of hGH, stimulated erythropoiesis in vitro in the presence of erythropoietin, as demonstrated by the increased activity of a cytosolic enzyme of the heme pathway (uroporphyrinogen I synthase). The results suggest that IGF-I could be involved in the regulation of erythroid differentiation.

COMPARATIVE EFFECTS OF HUMAN GROWTH HORMONE (GH) AND INSULIN-LIKE GROWTH FACTOR I/SOMATOMEDIN C (IGF I/ SMC) ON ERYTHROPOIETIC CULTURES FROM NORMAL CHILDREN

Androgen receptors in the neo-natal period: evidence for some “physiological” androgen resistance in male infants

The mechanism of androgen's action on cartilage growth plate is unknown. The aim of this study was to determine if receptors for androgen exist in dog and child epiphyseal growth cartilage. Cartilages were removed by microsurgical technique, from male dog bones, and at autopsy of a male newborn. 3H-DHT binding activity was evaluated in cytoplasmic and crude nuclear fractions of dog and child growth cartilage. Specifically bound 3H-DHT was determined by incubations of cytoplasmic or nuclear extracts with 3H-DHT (1-30 nM) alone or in the presence of an excess of unlabelled DHT to correct for non-specific binding. 3H-DHT specifically bound is 2 times higher in nuclear than in cytosolic fractions. Competition analysis revealed a high specificity of the binding component for DHT. Sucrose density gradient analysis of 3H-DHT labelled cytosol or nucleus from dog or child cartilage yielded one peak of radioactivity in the 4-S region. The high affinity, apparent specificity for DHT, and sedimentation analysis fulfill several of the criteria ascribed to a "receptor". We thus suggest that the macromolecular protein binding we describe in dog and child growth plate cartilage is an androgen receptor.

Mixed gonadal dysgenesis is characterised by female ductal differenciation in spite of presence of testicular tissue. Two cases of familial mixed gonadal dysgenesis are reported. The first patient had a female phenotype and presented at 13 years undeveloped breast, primary amenorrhea and enlarged clitoris. She had normal female external genitalia and a narrow vaginal introitus distinct from urethra. Normal vagina, uterus and tubes were opacified by external genitography. At laparotomy uterus appeared small but Falloppe tubes were normal. Streak gonad was found in the place of the right ovary and the site of the left gonad was occupied by an ovoid tumor which histologic examination occurred a gonadoblastoma with abnormal testicular tissue. The caryotype in blood culture was XY. The brother presented at birth hypospadias with bilateral cryptorchidism. He was 8 years when referred to the Paediatric Center because of ambiguous external genitalia. He had a male phenotype with labial fusion and fusiform urogenital sinus opening at base of phallus. Gynecography disclosed normal vagina and outset of uterus. Exploratory laparotomy revealed a right gonad: dysgenetic testis was found at biopsy. A next surgical laparotomy is programmed on opposite gonad. Caryotype was XY in blood culture. These reports may be classified in mixed gonadal dysgenesis group. Original findings = Same a−symetric testicular dysgenesis in two siblings, but with different phenotype. Its origin can be found in a structural abnormality of Y chromosome.

Several groups are still investigating an "in vitro" biological response of androgen action. The purpose of this study was to determine whether dihydrotestosterone (DHT) influences the secretion of collagen by cultured foreskin fibroblasts raised from patients with normal or undetectable DHT-binding activity (complete androgen resistance). Collagen synthesis and secretion were evaluated by 3H-proline incorporation in newly synthetized collagen by confluent fibroblasts alone, or in the presence of DHT (10−5 to -10−10M). Production of collagen is expressed as a ratio between tritiated OH-proline and OH-proline plus proline,within the cells or in the culture medium. These data show that : -in control cells,DHT significantly increases production of collagen released in the culture medium (p < 0.001), -In complete androgen resistance with no detectable androgen binding activity, DHT does not modify basal collagen synthesis. These results strongly suggest that this hormonal response is mediated via a DHT-receptor or is receptor-dependent. This could be used as a biological response of androgen action in patients with androgen resistance associated with a "post-receptor" defect.

The etiology of durable pubertal gynecomastia remains unexplained. Hormonal studies are controversial and receptor measurements inconclusive. A local conversion of androgens has been suggested. We have studied glandular and adipose fractions obtained from mastectomy of 3 adolescent boys. Microsomal suspension of the tissues was incubated with 14C testosterone or 3H-Δ4-androstenedione. Aromatization activity was measured by tritiated water formed during the incubation. Placental microsomes served as control. The identification of estrogen products was performed as follows: sodium partition, filtration on isomerase column, elution of the radioactivity corresponding to estrogens (E3) and finally gel chromatography of that extract. In placental microsomes, aromatase activity was: 0.66 pmoles/mg protein/min; in adipose tissues: 0.14 fmoles/mg protein/min; in glandular tissues: 0.13 fmoles/mg protein/min. We conclude that in vitro gynecomastic glandular tissue transforms androgen to the same extent as adipose tissue. An exaggerated local estrogen formation can most likely be excluded.

Case 1 : Born 1976, female sex assigned, Karine. Examination at age 16 months : arbiguous genitalia (Prader II), phallus 1,5 cm length, with erections, palpable gonads in inguinal area; genitography : rudimentary vagina. Laparotomy : no uterus, 2 testis with epididymis, normal histology. Caryotype XY. Testosterone raises from 14 to 203 ng% with HCG. At age 5, bilateral orchidectomy is performed and phalloplasty in order to suppress erections and to confirm female sex. At age 9, Karine is a well being little girl. Binding capacity of DHT receptors on sexual skin fibroblasts is 204 fentcmoles/mg DNA, kd : 0,8 nM. Case 2 : Born 1979, male sex assigned, Mickael.Micropenis 0,5 cm length. No circonferential forehead skin. XY caryotype. Testosterone raises from 16 to 65 ng% with HCG. No mullerian structure seen at genitography. Normal male urethre. With androgen therapy, penis size reaches 4 cm length with erections and lack of bone age excess of maturation. At age 6, orchidopexy. Fair male evolution. Before treatment, aged 2, binding capacity of DHT-receptors on sexual skin fibroblasts is 286 fentomoles/mg DNA, Kd : 0,65 nM (N : 400 to 800). At age 7, receptors level is 484 fentomoles/mg DNA, Kd : 0,33 nM on penis skin and 890 fentomoles/mg DNA, Kd : 0,54 nM on scrotal skin. Then, a partial receptor deficiency is expressed in different phenotypic way in these two siblings with XY caryotype since the one was maintened in the female sex and the other in the male sex. In this latter, normalization of external genitalia was obtained with long term androgen therapy which is explained by the remarkable raise of receptor-sites observed at 5 years intervall.

We studied the role of androgens on children erythropoietic precursors cells in culture. Cultures of normal marrow from surgical intervention (informed consent) were carried out using a miniaturized methyl-cellulose method in the presence of erythropoietin (11U/ml). Effects of testosterone (T), 5 -dihydrotestosterone (DHT) and nor-testosterone (nor-T) were evaluated on colony forming units erythroid (CFU-E) after 7days of incubation at 37°C CFU-E were quantified by scoring directly colonies and by a biochemical determination of the uroporphyrinogen I synthase activity (UROS). Results are given as number of CFU-E per 32.000 nucleated cells plated. UROS activity is expressed as pmoles of uroporphyrinogen formed per hour and per well. T,DHT and nor-T significantly stimulate children marrow progenitor cells in culture.Androgens act on both growth and maturation of CFU-E.A direct effect of androgens on erythroid differentiation (via an androgen receptor?) in children bone marrow culture may explain the efficiency of androgen treatment in aplastic anemia.

It has been suggested that rate of estrogen formation was higher in patients with Androgen Resistance Syndrome (ARS). This work was designed to find out if peripheral aromatase activity could be related to a defect in androgen action in prepubertal children with male pseudohermaphroditism. Fibroblast estrogen production was assayed by a highly specific enzymatic determination. Foreskin fibroblast strains were raised from 27 children with complete (C.ARS) or partial (P.ARS) androgen resistance as defined by dihydrotestosterone binding activity in cells. Results (mean ± SD) are expressed as pmoles estrone/mg proteins synthetized per day when cultured fibroblasts are incubated with Δ4-androstenedione. These results show that -peripheral aromatase activity is low before puberty -fibroblast estrogen synthesis is significantly increased in prepubertal children with partial or complete ARS. Our data suggest that low utilization of androgens by target cells stimulates the production of estrogen. Peripheral aromatase activity can thus be considered as a marker of androgen resistance in prepubertal children with male pseudohermaphroditism.

Longitudinal studies are very adequate to detect small individual and significant variations of plasmatic hormons. Thus we performed prolactin determination (PRL) in 12 boys and 9 girls who were patients, in the pediatric department of CHU St-Charles for non-endocrine chronic diseases. PRL determinations were done according to Reuter et al. (1975, Path. Biol. 23, 761-767) using commercial Kits (IRE-Fleury, Belgium) tested in our laboratory for specificity and reproductibility with NIH and MRC standards. In our hands, the “intra-assay” error was 5 %, the “inter-assay” error 10 %, and the equivalence was found to be 40 μI.U.= 1 ng NIH, VLS-1. In boys (n = 12) the mean PRL values as a function of Tanner's puberal stages were found to be (ng/ml) : stage I : 4.4±0.2; stage II : 3.3±0.2; stage III : 2.7±0.2; stage IV : 2.2±0.3; stage V : 4.8±0.2 Between stages II and IV, an important decrease of PRL was observed which was followed by an increase at the end of puberty, PRL becoming slightly higher than at stage I. This pattern could not be detected by a non-longitudinal study. In girls, PRL levels (ng/ml) increase progressively throughout puberty : stage I : 3.0±0.3; stage II : 3.7±0.2; stage III : 3.9±0.2; stage IV : 4.6±0.3; stage V : 6.0±0.4. These results agree with PRL levels observed in groups of prepuberal and postpuberal girls. Thus PRL variations during puberty are made obvious by longitudinal studies but their eventual implication in the maturation process of the hypothalamo-pituitary gonadal axis is still to be determined.

Previous studies in this laboratory have shown that human cultured skin fibroblasts have a receptor which binds dihydrotestosterone (DHT) with high affinity, high specificity but low capacity. Kd and Bmax values were similar in sex skin fibroblasts from male and female subjects but the Bmax of sex skin was higher than in non-sex skin. The present investigation was designed to find out whether fetal cells have DHT receptors, the age they can be detected and whether there are differences in Bmax between fibroblasts from sexual organs and non-sexual tissues. 1/ The youngest specimen studied was 8 wks: DHT receptors were found in fibroblasts from non-sex skin. 2/ A 10 wk fetus had a high level of DHT receptors in fibroblasts from the external genitalia. The value was similar to normal values for newborn skin. 3/ In 2 female fetuses there was greater DHT binding in fibroblasts from sex organ than in those from non-sexual skin. 4/ DHT receptors were present in fibroblasts from testis, from lung but not from intestine. 5/ In a mixture of epithelial cells and fibroblasts from one kidney, we found receptors with a low Bmax and a low Kd. 6/ 5α-reductase activity was detected in fibroblasts of a 10 wk old fetus; the levels being lower in non-sex skin than in sex organs.

ANDROGEN METABOLISM AND SPECIFIC 5α-DIHYDROTESTOSTERONE (DHT) RECEPTORS IN HUMAN AMNIOTIC FLUID FIBROBLASTS

Testosterone (T) and dihydrotestosterone (DHT) were found to bind to a specific protein in foreskin fibroblasts of a male pseudohermaphrodite with 5α-reductase deficiency. While maximum number of binding sites (B max) were similar for both androgens, the apparent dissociation constant (Kd) of the androgen receptor for T was greater than for DHT. In competition studies of 3H-T bound to the receptor with unlabeled T or DHT, the inhibitor constant (Ki) for T was two to three fold greater than the Kj for DHT. Also, the dissociation rate constant (kd) for 3H-T bound to the receptor was greater than for 3H-DHT (t 1/2 for T = 10 h and t 1/2 for DHT = 74.5 h). These results suggest that T may play a role in the sexual differentiation of patients with 5α-reductase deficiency. Their incomplete masculinization would be explained by the lower affinity and faster turnover rate of the T-receptor complex relative to the DHT-receptor complex.