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Biomedical subjects

B Terouanne

Publications and source records attributed to B Terouanne.

17 recordsLinked to original sources
PubMed2002

Disorders of androgen action.

Disorders of androgen action are the main cause of male pseudohermaphroditism and include 5alphaR deficiency and androgen receptor defects. 5alphaR deficiency is characterized by female genitalia with some degree of masculinization, clitoromegaly, and severely bifid scrotum corresponding to the so-called pseudovaginal perineoscrotal hypospadias. At the onset of puberty, increased muscle mass, development of pubic hair, and phallic growth are associated with the acquisition of male gender identity. Normal or increased levels of testosterone and an elevated testosterone-to-dihydrotestosterone ratio after human chorionic gonadotropin stimulation testing suggest 5alphareductase deficiency, and the diagnosis can be ascertained by identifying the mutation in the 5alphaR-2 gene. Whatever the patient's age at diagnosis, psychological evaluation with 5alphaRD is vital. Androgen receptor defects encompass two clinical expressions: the complete and partial androgen insensitivity syndromes. Complete androgen insensitivity syndrome should be suspected at birth in the presence of inguinal hernia in a girl without genital ambiguity. At puberty, the sign of alert is primary amenorrhea with normal female phenotype and harmonious mammary development but no pubic hair growth. Partial androgen insensitivity syndrome covers a wide spectrum of undervirilized phenotypes ranging from clitoromegaly at birth to infertile men. In all cases, complementary investigations should include plasma testosterone and luteinizing hormone as well as androgen-binding capacity in cultured genital skin fibroblasts. Diagnosis is confirmed by identification of the androgen receptor gene mutation. Although patients with complete androgen insensitivity syndrome are raised as females, patients with partial androgen insensitivity syndrome should be managed according to age at diagnosis, response to treatment with exogenous androgens, and the presence of an androgen gene mutation. Gonadectomy in complete androgen insensitivity syndrome should be performed before puberty, and androgen substitution may improve the development of external genitalia in some patients with partial androgen insensitivity syndrome. Psychological follow-up is necessary.

3-Oxo-5-alpha-Steroid 4-Dehydrogenase↗
PubMed2002

[The androgen receptor: molecular pathology].

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.

Active Transport, Cell Nucleus↗
PubMed2001

Environmental xenoestrogens, antiandrogens and disorders of male sexual differentiation.

Over the past 20 years, the documented increase in the disorders of male sexual differentiation, such as hypospadias, cryptorchidism, and micropenis, has led to the suspicion that environmental chemicals are detrimental to normal male genital development in utero. Male sexual differentiation is critically dependent on the normal action of androgens, and unbalanced androgen/estrogen ratios can disturb it. Environmental xenoestrogens (such as herbicides, pesticides, PCBs, plasticizers, and polystyrenes) that mimic estrogens or environmental antiandrogens (such as polyaromatic hydrocarbons, linuron, vinclozolin, and pp'DDE) that disturb endocrine balance, cause demasculinizing effects in the male foetus. These environmental chemicals are often referred to as endocrine disruptors: they are thought to mimic endogenous estrogens by entering the cell, binding to the receptor and activating transcription, they may also antagonize normal androgen action. We have established numerous cell lines to assess the estrogenicity and antiandrogenicity of compounds found in the environment and to identify new products present in wastewater effluents that are able to disrupt endocrine functions. Several cell lines responding to estrogens have been obtained in our group, including cells with different enzymatic equipment and cells expressing chimeric receptor or natural estrogen receptors alpha and beta. These cell lines have proved to be useful for assessing the biological activity of pesticides, fungicides, and chemicals found in plastic or discarded in the environment. In order to generate a powerful tool for the investigation of androgen action and the rapid screening of potential antagonists, we developed a new stable prostatic cell line. The PALM cell line is an original cellular model to characterize the response of hAR, and it provides an easy and rapid bioluminescent test to identify new antagonists. We also developed a model based on a fusion protein between the androgen receptor (AR) and the green fluorescent protein (GFP) to study the intracellular dynamics of AR. The GFP-AR model was applied to define the ability of several xenoestrogens and antiandrogens to inhibit the nuclear transfer of AR. The ubiquitous presence of endocrine disruptors in the environment and the increased incidence of neonatal genital malformation support the hypothesis that disturbed male sexual differentiation may in some cases be caused by increased exposure to environmental xenoestrogens and/or antiandrogens.

Androgen Antagonists↗
PubMed1997

Trafficking of the androgen receptor in living cells with fused green fluorescent protein-androgen receptor.

The trafficking of the androgen receptor (AR) in transfected cells was studied using a green fluorescent protein (GFP)-AR chimera. The reporter molecule GFP enabled the localization of AR in living cells with a good spatial and temporal resolution. After the construction of GFP-AR and verification of the size of the fusion protein produced, we demonstrated that GFP-AR conserves the functional properties of the AR: GFP-AR had the same androgen-binding affinity as AR, and GFP-AR efficiently transactivated an androgen-responsive gene in response to synthetic androgen at 30 degrees C. The fusion protein was first detected throughout the cytoplasm without hormone, fluorescence becoming nuclear rapidly after androgen incubation. This hormone dependence of AR trafficking was confirmed by the use of the mutant GFP-AR-del4, which lacked the androgen-binding function. The mutant was localized in the cytoplasm in the absence of hormone, but the distribution was not modified by androgen incubation. An ACAS 570 scanning laser cytometer was used to quantify fluorescence in a single living cell, first without and then with hormone. Different hormones and antihormones were tested to determine the dynamics of GFP-AR translocation into the nucleus. All the drugs used were able to induce nuclear translocation, and steady state level was rapidly attained within 1 h. The ratio of receptors in cytoplasmic and nuclear compartments was related to both affinity and concentration of ligand. The data from this follow-up study demonstrated for the first time the intracellular dynamics of the hormone-dependent trafficking of AR in a single living cell.

Androgens↗
PubMed1994

Hydroxytamoxifen induces a rapid and irreversible inactivation of an estrogenic response in an MCF-7-derived cell line.

The MVLN cell line was established in our laboratory from MCF-7 cells by stable transfection with the luciferase gene under the control of an estrogen-responsive element from the Xenopus vitellogenin A2 gene. This cell line allowed us to visualize the induction by hydroxytamoxifen of a heterogeneity in the cell population with regard to the expression of the luciferase gene. Treated cells lost their estradiol-inducible luciferase activity, progressively and irreversibly; the luciferase expression of 80% of the cells was irreversibly inactivated by a 12-day hydroxytamoxifen treatment. We showed that this inactivation process was specific for an estrogenic response and was mediated by the estrogen receptor. Tamoxifen itself gave rise to such an inactivation, whereas other compounds belonging to the triphenylethylenic family but differently substituted on the ethylenic carbon and the ICI 164,384 compound were not as efficient. This irreversible inactivation was accompanied by a sharp decrease in the luciferase mRNA level; however, the estrogen receptor function and the cellular transcriptional machinery were not affected by the treatment. Although this antiestrogen treatment neither affected the estrogen-dependent cell growth nor irreversibly inhibited the expression of the natural pS2 gene, these results highly suggest that long-term antiestrogen therapy may lead to some heterogeneity in tumor cells throughout the course of patient treatment.

Base Sequence↗
PubMed1992

A multiple luminescent procedure for the detection of different papillomaviruses on dot blots.

We developed a method based on the use of various luminescent systems for identification of several nucleic acid sequences on the same dot blots. In a simultaneous assay, the target DNA molecules were hybridized with different DNA probes. These probes were labelled with biotin or digoxigenin or directly coupled to enzymes such as glucose-6-phosphate dehydrogenase, peroxidase or alkaline phosphatase. After hybridization, these labels were detected by luminescent reactions using an amplified camera. Rapid detection and specific identification of pathogenic agents such as human papillomaviruses (HPV) could be performed in a single step by this procedure. Polymerase chain reaction was carried out using general primers and virus types were identified on dot blots using short HPV 11, 16 and 18 specific oligonucleotides.

Base Sequence↗
PubMed1989

[Use of a technic of bioluminescence for determining estrogens and LH during static and dynamic study of the ovary].

Estrogens and LH are necessary among other assays for ovulation diagnostic and ovarian monitoring during stimulations. They can be both measured with a method based on a bioluminescent reaction. This method measures, with a standard luminometer, the reduced NAD produced and accumulated by the reaction of two dehydrogenase enzymes: the estradiol dehydrogenase for direct assay of estrogens (estrone + estradiol), the glucose-6-phosphate dehydrogenase for gonadotrophin determination. These techniques can be applied to all biological fluids. They are simple and straightforward, do not need extraction and can be automated. Urinary assays are very useful in clinical practice to appreciate ovarian function, either during a spontaneous cycle or under stimulation, and are mandatory to decide the timing of hCG injection. The reported studies can be listed as follows: determination of preovulatory LH rise (LH greater than 10 Ul/g of creatinine) prior to embryo transfer after cryopreservation and thawing, detection of LH surges during ovarian stimulation, either premature surges causing premature luteinization, either normal surges when follicular maturation is adequate (296 cycles), confirmation of pituitary desensitization when using GnRH agonists (43 cycles), study of the initial stimulatory effect of GnRH agonists (13 patients). This effect can be responsible for the inadequate results obtained with the so-called "short protocol" in this experience when compared with the "long protocol" in the author's experience with compared with the "long protocol" (8 p. cent pregnancy rate per stimulation cycle versus 20 p. cent respectively, intrafollicular LH in 129 follicular fluids (86 with GnRH agonists and 43 without) has no correlation with the fecundability of the ovum. These results lead to extend bioluminescent techniques to the study of other parameters, and in particular FSH.

Buserelin↗
PubMed1989

[Bioluminescence assay of luteinizing hormone in plasma and urine].

The authors describe a bioluminescent immunoassay of LH in plasma and urine. It uses two monoclonal antibodies, one is labelled with glucose-6-phosphate-dehydrogenase, the other one is coimmobilized together with bioluminescent enzymes from marine bacteria on the same adsorbent (Sepharose). This assay can be performed directly on 20 microliters plasma or 10 microliters urine. The protocol is very fast, no separation step is required to remove the excess labeled antibodies. The inhibitory effect of the biological sample on luminescent reaction is determined by adding NADH to the assay tubes. The working range of this assay is 3 to 300 Ul/l, with a sensitivity of detection of 0.5 Ul/l. Recovery, linearity, within and between assay precision were evaluated and appeared to be satisfactory. The authors have observed a good correlation between results obtained with our method and with radioimmunoassay.

Female↗
PubMed1985

[Chemiluminescence and bioluminescence in clinical analysis. Perspectives of development].

Until now, the methods of analysis by luminescence have not developed greatly in clinical biology because of the instability of the reagents and the absence of a single protocol applicable to a sufficiently wide range of assays. For this reason, the application has been limited to several assays using a specific bioluminescence reaction i.e., the ATP assay (for the measurement of bacterial contamination), and several assays which evaluate NADH production (bile acids, triglycerides, lactate). Research on assay methods that are very specific and very sensitive and do not use radioactive molecules has led to, among other things, the development of new techniques using luminescence reactions. These reactions are of two types: chemiluminescence reactions, which use molecules (luminol, dioxetane, derivatives of acridine and imidazol) that lead to an emission of light under the action of various chemical and enzymatic compounds; bioluminescence reactions produced by specific enzymatic systems (luciferases). Chemiluminescence reactions are used in several immunometric assays. The chemiluminescent marker fixed to the antigen or antibody has a low molecular weight: for this reason it does not lead to a significant modification of immunoreactivity and it is highly stable. Moreover, in certain cases it is possible to use particular properties of these markers (energy transfer, modification of the efficiency of the luminescent reaction) to carry out homogeneous immunometric assays requiring no separation step. Bioluminescence reactions have been less developed, because of the instability of enzymatic systems. However, these reactions are by far the most sensitive of all the detection techniques currently used.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenosine Triphosphate↗
PubMed1983

Centrifugal analyzer used for enzyme immunoassay of progesterone and choriomammotropin.

Recently we developed an enzyme immunoassay involving the use of steroid delta-isomerase (EC 5.3.3.1) as enzyme label and exclusion-affinity chromatography for rapid separation of free antigen-enzyme conjugate that bound to antibodies (J. Immunol. Methods 35: 267-284, 1980). Here we describe an automated version of this procedure, for immunoassay of progesterone and choriomammotropin (human placental lactogen) in serum with the use of a centrifugal analyzer. After incubation, suitable dilutions of sera or extract plus antiserum, conjugate, and double antibody were filtered on an estradiol affinity gel-filtration column; the enzyme activity of the filtrates was determined with the centrifugal analyzer. Results correlate well with those obtained by radioimmunoassay: r (progesterone) = 0.980, r (choriomammotropin) = 0.940. The within-run and between-run precision, specificity, sensitivity, accuracy, and speed of this system make it a useful tool for immunoassay.

Autoanalysis↗
PubMed1980

Coupling of delta 5,3-ketosteroid isomerase to human placental lactogen with intermolecular disulfide bond formation. Use of this conjugate for a sensitive enzyme immunoassay.

An immunoassay for human placental lactogen (HPL) has been developed with the use of hormone labelled with delta 5,3-ketosteroid isomerase. The enzyme is coupled to the antigen via an intermolecular disulfide interchange reaction. HPL-enzyme conjugates are prepared in high yield and purified by affinity chromatography. An enzyme immunoassay using these hormone-enzyme derivatives is compared with radioimmunoassay for determination of HPL in serum.

Animals↗
PubMed1980

Affinity chromatography: a tool for enzyme immunoassay.

Affinity chromatography has been widely used for enzyme purification and for the preparation of antigen-enzyme conjugates. Most enzyme immunoassay use immunoadsorbent (specific or double insolubilized antibodies) which makes automation difficult as centrifugation and washing steps are necessary. We propose affinity chromatography for rapid separation of antigen-enzyme bound to antibodies from free conjugate and its application in enzyme immunoassay. We describe enzyme immunoassays for human placental lactogen and progesterone which use delta 5,3-ketosteroid isomerase as marker and depend upon retention by specific absorbent of enzyme conjugate not bound to antibodies.

Adsorption↗
PubMed1977

Characterisation of an associate 17-beta-hydroxysteroid dehydrogenase activity and affinity labelling of the 3-alpha-hydroxysteroid dehydrogenase of Pseudomonas testosteroni.

The 3-alpha-hydroxysteroid dehydrogenase and the 3-beta-hydroxysteroid dehydrogenase of Pseudomonas testosteroni were purified to homogeneity by polyaerylamide gel electrophoresis using the following stages: DEAE cellulose chromatography, affinity chromatography on oestrone-aminocaproate sepharose and Sephadex gel filtration. The pure 3-alpha-hydroxysteroid dehydrogenase was completely devoid of 3-beta-hydroxysteroid dehydrogenase activity but could oxidize estradiol 17-beta at an appreciable rate. This activity accounts for about 40 per cent of the total 17-beta-estradiol dehydrogenase of the crude bacterial extract. Affinity labelling of pure 3-alpha-hydroxysteroid dehydrogenase was carried out using 5-beta-pregnane 3,20-dione-12-alpha-iodoacetate and 5-alpha-androstane 3-one-17-beta-bromoacetate. With both reagents, inactivation was obtained only in the presence of coenzyme, the substrate protected against inactivation and the enzyme was fully inhibited with covalent binding of 1 mole of reagent per mole of subunit suggesting an active site directed inhibition. Histidine and methionine were identified as the labelled aminoacid residues.

17-Hydroxysteroid Dehydrogenases↗
PubMedDate not supplied

Disorders linked to insufficient androgen action in male children.

Virilization of the external genitalia in the male fetus requires testosterone and dihydrotestosterone (DHT), which is formed from testosterone by the action of the enzyme, 5alpha-reductase type 2 (5alphaR-2). Mediation of the effects of both testosterone and DHT requires a functional androgen receptor (AR) located in the cytoplasmic compartment of target cells. DHT (or testosterone) binding induces a conformational change which facilitates AR nuclear transport, phosphorylation and dimerization, ultimately regulating of the rate of transcription of androgen-dependent genes. Any event which impairs DHT formation (mutation within the 5alphaR-2 gene or 5alphaR-2 inhibitors) or normal function of the AR (mutation in the AR gene, antiandrogens) may result in insufficient androgen action in the male fetus and in subsequent undervirilization in the newborn. Hypospadias may be due to a defect in androgen action due to mutation of the 5alphaR-2 or of the AR gene. Mutation of unidentified genes is likely to underlie this displacement of the urethral meatus from the tip to the ventral side of the phallus. An aetiological role for environmental chemical products has been postulated, since ethnic as well as geographical differences in the incidence of hypospadias have been noted. Increasing evidence has been gathered indicating that widely used industrial and agricultural chemicals have deleterious effects on normal male sexual differentiation. Cryptorchidism and micropenis may represent an intersex phenotype, even if they are isolated. Aetiological factors include 5alphaR-2 gene mutation, AR gene mutation or environmental hormonal disruptors. In conclusion, several phenotypes have been attributed to insufficient androgen action during fetal life. Whereas mutations in the 5alphaR-2 gene and AR gene are natural, attention should be focused on environmental endocrine disruptors that are able to mimic steroid 5alpha-reductase deficiency or partial androgen insensitivity syndrome.

3-Oxo-5-alpha-Steroid 4-Dehydrogenase↗
PubMedDate not supplied

[Androgen-independent prostate carcinoma and androgen-receptor: recent progress in molecular genetics].

Prostate cancer is an androgen-dependent tumor which presents an androgen-independent regrowth after clinical regression in response to antiandrogen treatment. Four hypotheses have been developed to understand how androgen signal transduction pathway mediate androgen-independent tumor progression: over expression of the wild-type androgen-receptor gene, androgen-receptor gene mutation, excessive recruitment of transcriptional co-activator ARA-70 and a cross-talk between the androgen-receptor and the growth factor receptor pathways. In this work, C. Sawyers's group elegantly demonstrates, in LAPC-4 androgen-independent prostate cancer sublines, that forced hyperexpression of HER-2/Neu receptor tyrosine kinase allowed androgen-independent growth, that HER-2/Neu activated the androgen-receptor pathway in the absence of androgens and synergized with low levels of androgen to superactivate the pathway. These important data could have therapeutic implications for the management of androgen-independent prostate cancer.

Disease Progression↗