Inserm/Université U 845

- Physiopathologie des hormones de la famille PRL/GH : approches transversales -

Réponses affichées : 31

Lien PubMed New concepts in prolactin biology
BERNICHTEIN S, TOURAINE P, GOFFIN V
2010 - J Endocrinol 206(1):1-11
Human prolactin (PRL) is currently viewed as a hormone of pituitary origin, whose production (i.e. serum levels) is controlled by dopamine, whose biological actions relate exclusively to lactation and reproductive functions, for which any genetic disorder is yet to be identified, and whose unique associated pathology is hyperprolactinemia. Both experimental studies and human sample/cohort-based investigations performed during the past decade have considerably widened our perception of PRL biology: i) there are now strong epidemiological arguments supporting the fact that circulating PRL is a risk factor for breast cancer, ii) in addition to the endocrine hormone, locally produced PRL has been documented in several human tissues; there is increasing evidence supporting the tumor growth potency of local PRL, acting via autocrine/paracrine mechanisms, in both rodent models, and human breast and prostate tumors, iii) the first functional germinal polymorphisms of the PRL receptor were recently identified in patients presenting with breast tumors, which involve single amino acid substitution variants exhibiting constitutive activity, iv) human PRL analogs have been engineered, which were shown in experimental models to down-regulate the effects triggered by local PRL (competitive antagonism) or by the constitutively active receptor variants (inverse agonism). The aim of this review is to discuss these novel concepts in PRL biology, including their potential pathophysiological outcomes.
Unité(s) : U845 (VG)
 
Lien PubMed Fertility in Women with Nonclassical Congenital Adrenal Hyperplasia due to 21-Hydroxylase Deficiency
BIDET M, BELLANNE-CHANTELOT C, GALAND-PORTIER MB, GOLMARD JL, TARDY V, MOREL Y, CLAUIN S, COUSSIEU C, BOUDOU P, MOWZOWICZ I, BACHELOT A, TOURAINE P, KUTTENN F
2010 - J Clin Endocrinol Metab 95(3):1182-90
Objective: In contrast to subfertility often reported in women suffering from the classical form of congenital adrenal hyperplasia (CAH) due to 21-hydroxylase deficiency, fertility in nonclassical CAH (NC-CAH) has been rarely studied. Our objective was to evaluate fertility in NC-CAH women. Material and Methods: We studied 190 NC-CAH women (161 probands + 29 first degree relatives). Only 20 probands had consulted for infertility (12%), either alone or associated with hirsutism or menstrual cycle disorders. The diagnosis was established on post-ACTH 17-hydroxyprogesterone 10 ng/ml or greater and further characterized by CYP21A2 gene analysis. Results: Ninety-five of the 190 women wanted pregnancy (aged 26.7 +/- 8.9 yr); 187 pregnancies occurred in 85 women, which resulted in 141 births in 82 of them. Ninety-nine pregnancies (52.9%) occurred before the diagnosis of NC-CAH (96 spontaneously and three with ovulation inducers) whereas 98 occurred after diagnosis (11 spontaneously and 77 with hydrocortisone treatment); 83% of pregnancies were obtained within 1 yr. The rate of miscarriages was 6.5% for pregnancies obtained with glucocorticoid treatment vs. 26.3% without. Two of the 141 infants (1.5%) were born with classical CAH. Conclusion: Subfertility is mild in NC-CAH. However, the rate of miscarriages is lower in pregnancies occurring with glucocorticoid treatment and argues for treating NC-CAH women wanting pregnancy. In addition, considering the high rate of heterozygotes for CYP21A2 mutations in the general population, it is essential to genotype the partner of patients with a severe mutation to predict the risk of classical CAH and offer genetic counseling.
Unité(s) : U845 (VG)
 
Lien PubMed Crystal structure of an affinity-matured prolactin complexed to its dimerized receptor reveals the topology of hormone binding site 2
BROUTIN I, JOMAIN JB, TALLET E, VAN AGTHOVEN J, RAYNAL B, HOOS S, KRAGELUND BB, KELLY PA, DUCRUIX A, ENGLAND P, GOFFIN V
2010 - J Biol Chem 285(11):8422-33
We report the first crystal structure of a 1:2 hormone-receptor complex that involves prolactin (PRL) as the ligand, at 3.8 A resolution. Stable ternary complexes were obtained by generating affinity-matured PRL variants harboring an N-terminal tail from ovine placental lactogen, a closely-related PRL receptor (PRLR) ligand. This structure allows to draw up an exhaustive inventory of the residues involved at PRL/PRLR site 2 interface, consistent with all previously reported site-directed mutagenesis data. This description allows to propose an interaction model involving three structural components of PRL site 2 ("three pin plug"): the conserved glycine 129 of helix alpha3, the hydrogen bond network involving surrounding residues (glycine cavity), and the N-terminus. The model provides a molecular basis for the properties of the different PRL analogs designed to date, including PRLR antagonists. Finally, comparison of our 1:2 PRL/PRLR2 structure with those of free PRL and its 1:1 complex indicates that the structure of PRL undergoes significant changes when binding the first, but not the second receptor. This suggests that the second PRLR moiety adapts to the 1:1 complex rather than the opposite. In conclusion, this structure will be a useful guiding tool for further investigations of the molecular mechanisms involved in PRLR dimerization and activation, as well as for the optimization of PRLR antagonists, an emerging class of compounds with high therapeutic potential against breast and prostate cancer.
Unité(s) : U845 (VG)
 
Lien PubMed Characterization of two constitutively active prolactin receptor variants in a cohort of 95 women with multiple breast fibroadenomas
COURTILLOT C, CHAKHTOURA Z, BOGORAD R, GENESTIE C, BERNICHTEIN S, BADACHI Y, JANAUD G, AKAKPO JP, BACHELOT A, KUTTENN F, GOFFIN V, TOURAINE P
2010 - J Clin Endocrinol Metab 95(1):271-9
BACKGROUND: The role of prolactin (PRL) and its receptor (hPRLR) in promoting breast tumors is debated. We recently identified a gain-of-function hPRLR variant (I146L) in four women with multiple breast fibroadenomas (MFA) and no control subject. OBJECTIVES: The specific aims were to describe this cohort of women presenting with MFA to identify and functionally characterize germline variants of hPRL/hPRLR genes and compare phenotypes of all patients. DESIGN: Ninety-five patients prospectively underwent clinical examination, breast ultrasonography, magnetic resonance imaging, and hormonal evaluation of gonadal and lactotrope functions. We analyzed hPRL/hPRLR coding sequences and made comparisons with a control population of 194 women. Functional characterization of hPRLR variants was performed. Pathology and immunochemistry were systematically carried out after surgical removal of tumors. RESULTS: One third of patients had a family history of breast disease. No hormonal imbalance was observed, except 30.7% of explosive stimulated PRL. Prolactin receptor variants were identified in exon 5 (I76V: 10 patients, eight controls) and exon 10 (one patient, no control). Both I146L and I76V variants exhibited constitutive activity. Pathology showed common fibroadenomas and identified six benign phyllodes tumors. Estrogen and progesterone receptors were detected in 85 and 98% of samples, respectively. Ki-67 median staining was less than 5%. No phenotypic difference was observed between carriers and noncarriers of either hPRLR variant. CONCLUSION: We present the largest population with MFA ever described, 15% of which had a hPRLR exhibiting basal activity in vitro. This questions the involvement of the hPRLR in MFA etiology and the potential relevance of therapeutic inhibition of PRLR signaling in patients.
Unité(s) : U845 (VG)
 
Lien PubMed Prolactin and human tumorogenesis
FERNANDEZ I, TOURAINE P, GOFFIN V
2010 - J Neuroendocrinol 22(7):771-77
Summary The involvement of prolactin in human tumorogenesis has been long debated. The reason is that evidence supporting the role of circulating prolactin in promoting breast cancer was mainly obtained using rodent models, while most of the investigations performed in human species in the eighties have remained inconclusive. Things have started to change, as two alternative mechanisms of prolactin actions in tumor growth have emerged since the beginning of the 21(st) century. The first involves locally-produced prolactin, which acts by an autocrine/paracrine mechanism. Genetically-modified mouse models have demonstrated the tumorigenic potential of local prolactin on the prostate and the mammary gland, and arguments are now emerging in humans also. The second mechanism involves genetic variants of the receptor. While no genetic disorder has been reported for prolactin or its receptor, a variant of the prolactin receptor exhibiting constitutive activity has been recently identified in patients presenting with breast tumors, suggesting that sustained prolactin signalling may participate in breast tumorogenesis. Recent data regarding these two non classical mechanisms of prolactin action are discussed. Finally, we address the question of their inhibition in future cancer therapy, both in light of other findings that have revealed novel actions of prolactin in breast cancer cells, and with respect to the compounds currently available to target prolactin receptor signalling.
Unité(s) : U845 (VG)
 
Lien PubMed Mind the (gender) gap: does prolactin exert gender and/or site-specific effects on the human hair follicle?
LANGAN EA, RAMOT Y, GOFFIN V, GRIFFITHS CE, FOITZIK K, PAUS R
2010 - J Invest Dermatol 130(3):886-91
Unité(s) : U845 (VG)
 
Lien PubMed FOXL2 mutations lead to different ovarian phenotypes in BPES patients: Case Report
MEDURI G, BACHELOT A, DUFLOS C, BSTANDIG B, POIROT C, GENESTIE C, VEITIA R, DE BAERE E, TOURAINE P
2010 - Hum Reprod 25(1):235-43
FOXL2 mutations cause the autosomal dominant Blepharophimosis-ptosis-epicanthus inversus syndrome (BPES) that may be associated with premature ovarian failure (POF). However, little is known about the molecular mechanisms of FOXL2 actions in the human ovary. We conducted an extensive clinical, hormonal and ovarian histological study in two patients carrying a FOXL2 mutation associated with the typical eyelid malformations and infertility. This observational study was conducted at referral centres for POF. Histological and immunohistological studies were conducted on ovarian biopsies from two women with POF carrying a FOXL2 mutation resulting in putative polyalanine expansions of the protein. Abnormalities similar to those observed in mice with FOXL2 gene inactivation were present in the first patient's ovary, although the ovarian histology of the second patient was apparently normal. Different ovarian phenotypes, follicular defects and distribution of FOXL2 protein were observed in two patients carrying a FOXL2 mutation.
Unité(s) : Endocrinologie Pédiatrique et Gynécologie, U845 (VG)
 
Lien PubMed Prolactin--a novel neuroendocrine regulator of human keratin expression in situ
RAMOT Y, BIRO T, TIEDE S, TOTH BI, LANGAN EA, SUGAWARA K, FOITZIK K, INGBER A, GOFFIN V, LANGBEIN L, PAUS R
2010 - FASEB J 24(6):1768-79
The controls of human keratin expression in situ remain to be fully elucidated. Here, we have investigated the effects of the neurohormone prolactin (PRL) on keratin expression in a physiologically and clinically relevant test system: organ-cultured normal human hair follicles (HFs). Not only do HFs express a wide range of keratins, but they are also a source and target of PRL. Microarray analysis revealed that PRL differentially regulated a defined subset of keratins and keratin-associated proteins. Quantitative immunohistomorphometry and quantitative PCR confirmed that PRL up-regulated expression of keratins K5 and K14 and the epithelial stem cell-associated keratins K15 and K19 in organ-cultured HFs and/or isolated HF keratinocytes. PRL also up-regulated K15 promoter activity and K15 protein expression in situ, whereas it inhibited K6 and K31 expression. These regulatory effects were reversed by a pure competitive PRL receptor antagonist. Antagonist alone also modulated keratin expression, suggesting that "tonic stimulation" by endogenous PRL is required for normal expression levels of selected keratins. Therefore, our study identifies PRL as a major, clinically relevant, novel neuroendocrine regulator of both human keratin expression and human epithelial stem cell biology in situ.
Unité(s) : U845 (VG)
 
Lien PubMed Phenotyping and genetic studies of 357 consecutive patients presenting with premature ovarian failure
BACHELOT A, ROUXEL A, MASSIN N, DULON J, COURTILLOT C, MATUCHANSKY C, BADACHI Y, FORTIN A, PANIEL B, LECURU F, LEFRERE-BELDA MA, CONSTANCIS E, THIBAULT E, MEDURI G, GUIOCHON-MANTEL A, MISRAHI M, KUTTENN F, TOURAINE P
2009 - Eur J Endocrinol 161(1):179-87
OBJECTIVE: Premature ovarian failure (POF) encompasses a heterogeneous spectrum of conditions, with phenotypic variability among patients. The etiology of POF remains unknown in most cases. We performed a global phenotyping of POF women with the aim of better orienting attempts at an etiological diagnosis. DESIGN AND METHODS: We performed a mixed retrospective and prospective study of clinical, biological, histological, morphological, and genetic data relating to 357 consecutive POF patients between 1997 and 2008. The study was conducted at a reproductive endocrinology referral center. RESULTS: Seventy-six percent of the patients presented with normal puberty and secondary amenorrhea. Family history was present in 14% of the patients, clinical and/or biological autoimmunity in 14.3%. Fifty-six women had a fluctuating form of POF. The presence of follicles was suggested at ultrasonography in 50% of the patients, and observed in 29% at histology; the negative predictive value of the presence of follicles at ultrasonography was 77%. Bone mineral density alterations were found in 58% of the women. Eight patients had X chromosomal abnormalities other than Turner's syndrome, eight other patients evidenced FMR1 pre-mutation. Two other patients had autoimmune polyendocrine syndrome type 2 and 1. CONCLUSION: A genetic cause of POF was identified in 25 patients, i.e. 7% of the whole cohort. POF etiology remains most often undiscovered. Novel strategies of POF phenotyping are in such content mandatory to improve the rate of POF patients for whom etiology is identified.
Unité(s) : Pédiatrie Générale, Radiologie Adulte, U845 (VG)
 
Lien PubMed Adrenal hyperplasia and tumours in mice in connection with aberrant pituitary-gonadal function
BERNICHTEIN S, PELTOKETO H, HUHTANIEMI I
2009 - Mol Cell Endocrinol 300(1-2):164-168
Gonadectomy induces in certain inbred stains of mice adrenal hyperplasia and tumorigenesis, originating from the putative subcapsular stem/progenitor cell layer. This response is apparently triggered by the elevated post-gonadectomy levels of luteinising hormone (LH), followed by ectopic upregulation of adrenal LH/chorionic gonadotrophin (CG) receptors (Lhcgr). The clear strain dependence of this adrenal response to gonadectomy prompted us to study its genetic basis. Tumorigenic DBA/2J and non-tumorigenic C57BL/6J mice, as well as their F2 and backcrosses, were studied by whole genome linkage analysis. Gonadectomy induced similar upregulation of adrenal Lhcgr in both parental strains and their crosses, irrespective of the tumour status, indicating that ectopic expression of this receptor is not the immediate cause of tumours. Linkage analysis revealed one major significant quantitative trait locus (QTL) for the tumorigenesis on chromosome 8, modulated by epistasis with another QTL on chromosome 18. Hence, post-gonadectomy adrenal tumorigenesis in DBA/2J mice is a dominant trait, not a direct consequence of adrenal Lhcgr expression, and is driven by a complex genetic architecture. A promising candidate gene in the tumorigenesis linkage region is Sfrp1 (secreted frizzled-related protein 1), a tumour suppressor gene, which was down-regulated in the neoplastic tissue. Our findings may have relevance to the human pathogenesis of macronodular adrenal hyperplasia and postmenopausal adrenocortical tumours. A distinctly different adrenal response was observed in TG mice overexpressing LH or CG, or a constitutively activated form of the follicle-stimulating hormone receptor (Fshr). These mice developed perimedullary hyperlasia of foamy multinucleated cells, reminding of macrophages and filled with lipofuscin. Similar response was observed in TG mice overexpressing aromatase (CYP19). The cause of this response is not related to direct LH/CG action, but merely to adrenal response to chronically elevated oestrogen levels. This phenotype is reminiscent of the rare 'black adenomas' of the human adrenal cortex.
Unité(s) : U845 (VG)
 
Lien PubMed Stromal growth and epithelial cell proliferation in ventral prostates of liver X receptor knockout mice
KIM HJ, ANDERSSON LC, BOUTON D, WARNER M, GUSTAFSSON JA
2009 - Proc Natl Acad Sci U S A 106(2):558-63
With specific liver X receptor alpha and beta (LXRalpha and LXRbeta) antibodies, we found that LXRalpha is strongly expressed in the luminal and basal cells of prostatic epithelium. The ventral prostates (VP) of LXRalpha(-/-) mice are characterized by the presence of smooth-muscle actin-positive stromal overgrowth around the prostatic ducts and by numerous fibrous nodules pushing into the ducts and causing obstruction, so that most of the ducts were extremely dilated. BrdU labeling and Ki67 staining revealed epithelial and stromal proliferation in the fibrous nodules. However, the dense stroma surrounding the ducts was not positive for proliferation markers. There was no detectable difference between WT and LXRalpha(-/-) mice VP in the expression of the androgen receptor, but there was an increase in nuclear expression of Snail and Smad 2/3, indicating enhanced TGF-beta signaling. Upon treatment of WT mice for 3 months with the LXR agonist T2320 or for 3 weeks with beta-sitosterol, LXRalpha was downregulated, and a VP phenotype similar to that of LXRalpha(-/-) mice resulted. We conclude that in rodents, LXRalpha seems to control VP stromal growth and that LXRalpha(-/-) mice may be a useful model to study prostatic stromal hyperplasia. Because LXRalpha is expressed in the epithelium, the excessive stromal growth in LXRalpha(-/-) mice indicates that LXRalpha is essential for epithelial stromal communication.
Unité(s) : U845 (VG)
 
Lien PubMed Prolactin does not affect human platelet aggregation or secretion
REUWER AQ, NIEUWLAND R, FERNANDEZ I, GOFFIN V, VAN TIEL CM, SCHAAP MC, BERCKMANS RJ, KASTELEIN JJ, TWICKLER MT
2009 - Thromb Haemost 101(6):1119-27
Platelets play an important role in the development of plaque formation and in the events after rupture of the atherosclerotic plaque, leading to atherothrombosis. Multiple hormones, either in excess or when deficient, are involved in the development of atherothrombotic disease, but, to which extent such hormones affect platelet function, is still controversial. It was the objective of this study to assess the ability of the pituitary hormone prolactin to affect platelet functions. Venous blood was collected from six healthy males. Platelet activation was studied by (i) flow cytometry in whole blood (exposure of P-selectin as a measure of platelet secretion, and binding of PAC-1 as a measure of ligand-binding conformation of alpha(IIb)beta(3)), and by (ii) optical aggregation and whole blood aggregation. All studies were performed without and with exposure to several concentrations of ADP (0.1, 0.5 and 1.0 microM) and prolactin (50 and 1,000 microg/l). The presence of the prolactin receptor was investigated by Western blot and flow cytometry. In response to either 50 or 1,000 microg/l prolactin, no evidence of platelet activation or aggregation was found. In addition, ADP-induced platelet activation or aggregation was not enhanced by prolactin. Finally, prolactin receptors could not be detected on the surface of platelets. The present data indicate that prolactin does not directly modulate platelet function.
Unité(s) : U845 (VG)
 
Lien PubMed Prolactin Levels and the Risk of Future Coronary Artery Disease in Apparently Healthy Men and Women
REUWER AQ, TWICKLER MT, HUTTEN BA, MOLEMA FW, WAREHAM NJ, DALLINGA-THIE GM, BOGORAD RL, GOFFIN V, SMINK-BOL M, KASTELEIN JJ, BOEKHOLDT SM, KHAW KT
2009 - Circ Cardiovasc Genet 2(4):389-395
BACKGROUND: Prolactin is increasingly recognized to play a stimulatory role in the inflammatory response. Because inflammation is considered of crucial importance in the development of atherosclerosis, we aimed to evaluate whether prolactin levels are associated with the occurrence of coronary artery disease (CAD). METHODS AND RESULTS: We performed a nested case-control study in the prospective EPIC-Norfolk cohort. Cases were apparently healthy men and women, aged 45 to 79 years, who developed fatal or nonfatal CAD (n=882). Controls remained free of CAD (n=1490). Overall, systemic prolactin levels did not differ between cases and controls, and people in the highest prolactin tertile did not have a significantly increased risk of developing future CAD (in men, odds ratio, 1.21; 95% CI, 0.92 to 1.61; in women, odds ratio, 1.12; 95% CI, 0.76 to 1.64). However, in a separate immunohistochemical study, the presence of prolactin receptors could be demonstrated in postmortem human coronary artery plaques (preliminary data). CONCLUSIONS: Elevated systemic prolactin levels do not predict CAD in the general population. However, prolactin receptors were found in human coronary artery plaques. This observation may indicate a role of prolactin within atherosclerotic plaques. More studies are needed to define the possible role of prolactin in atherosclerotic plaque development.
Unité(s) : U845 (VG)
 
  Is the adrenal cortex a target for gonadotropins ?
BERNICHTEIN S, ALEVIZAKI M, HUHTANIEMI I
2008 - Trends Endocrinol. Metab. 19(7):231-238
The human adrenal cortex expresses low levels of luteinizing hormone/chorionic gonadotropin receptors (LHCGR), a characteristic gonad-specific G-protein coupled receptor (GPCR). LHCGR levels increase in the adrenal cortex after exposure to chronically elevated gonadotropins (e.g. after gonadectomy). In fact, heightened ectopic LHCGR levels are observed in a subclass of human adrenocortical tumors, and gonadotropin-responsive adrenocortical hyperplasia and tumors occur in several animal species. These findings suggest that adrenocortical responsiveness to LH/CG might be a physiological phenomenon that is amplified in the presence of elevated gonadotropin levels. Such increased gonadotropin action can induce pathologies ranging from adrenocorticotropic hormone (ACTH)-independent Cushing syndrome to malignant adrenal tumors. The authors review the current information on adrenocortical responses to gonadotropins in experimental animals and humans.
Unité(s) : U845 (VG)
 
  Identification of a gain-of-function mutation of the prolactin receptor in women with benign breast tumors
BOGORAD RL, COURTILLOT C, MESTAYER C, BERNICHTEIN S, HARUTYUNYAN L, JOMAIN JB, BACHELOT A, KUTTENN F, KELLY PA, GOFFIN V, TOURAINE P
2008 - Proc. Natl. Acad. Sci. USA 105(38):14533-14538
There is currently no known genetic disease linked to prolactin (Prl) or its receptor (PrlR) in humans. Given the essential role of this hormonal system in breast physiology, we reasoned that genetic anomalies of Prl/PrlR genes may be related to the occurrence of breast diseases with high proliferative potential. Multiple fibroadenomas (MFA) are benign breast tumors which appear most frequently in young women, including at puberty, when Prl has well-recognized proliferative actions on the breast. In a prospective study involving 74 MFA patients and 170 control subjects, we identified four patients harboring a heterozygous single nucleotide polymorphism in exon 6 of the PrlR gene, encoding Ile(146)-->Leu substitution in its extracellular domain. This sole substitution was sufficient to confer constitutive activity to the receptor variant (PrlR(I146L)), as assessed in three reconstituted cell models (Ba/F3, HEK293 and MCF-7 cells) by Prl-independent (i) PrlR tyrosine phosphorylation, (ii) activation of signal transducer and activator of transcription 5 (STAT5) signaling, (iii) transcriptional activity toward a Prl-responsive reporter gene, and (iv) cell proliferation and protection from cell death. Constitutive activity of PrlR(I146L) in the breast sample from a patient was supported by increased STAT5 signaling. This is a unique description of a functional mutation of the PrlR associated with a human disease. Hallmarks of constitutive activity were all reversed by a specific PrlR antagonist, which opens potential therapeutic approaches for MFA, or any other disease that could be associated with this mutation in future.
Unité(s) : U845 (VG), Endocrinologie Pédiatrique et Gynécologie
 
  Differences in prolactin receptor (PRLR) in mouse and human fallopian tubes: evidence for multiple regulatory mechanisms controlling PRLR isoform expression in mice
SHAO R, NUTU M, WEIJDEGARD B, EGECIOGLU E, FERNANDEZ-RODRIGUEZ J, TALLET E, GOFFIN V, LING C, BILLIG H
2008 - Biol. Reprod. 79(4):748-757
The anterior pituitary-derived hormone prolactin (PRL) signals through the PRL receptor (PRLR) and is important for female reproductive function in mammals. In contrast to the extensive studies of PRLR expression and regulation in human and mouse ovary and uterus, the mechanisms controlling the regulation of PRLR isoform expression in the fallopian tube are poorly understood. Because dynamic interaction of hormonal signaling in gonadal tissue and the pituitary or in gonadal tissues themselves in mammals suggests endocrine or paracrine regulation of PRLR expression, we questioned whether differential regulation of PRLR isoforms by PRL ovarian-derived estrogen (E(2)) and progesterone (P(4)) exists in the fallopian tube and pituitary of prepubertal female mice. Western blot analysis showed distinct molecular separation of PRLR isoforms in mouse and human fallopian tubes, and cellular localization was found in mouse and human tubal epithelia but not in mouse tubal smooth muscle cells. These data support the concept of an isoform- and cell type-specific expression of PRLR in human and mouse fallopian tubes. Moreover, expression of the long form of PRLR decreased after PRL treatment and increased after blockage of endogenous PRL secretion by bromocriptine (an inhibitor of PRL secretion) in a time-dependent manner in mouse fallopian tube. The opposite regulation was observed in the pituitary. Treatment with exogenous E(2) or P(4) led to changes in PRLR expression in the fallopian tube similar to those of PRL treatment. However, E(2) and P(4) did not affect PRLR expression in the pituitary. Estrogen had no effect on the long form of PRLR expression, whereas P(4) regulated the long form of PRLR in the fallopian tube, as did PRL. Taken together, the data from our comparative study provide evidence that PRLR can be regulated by an interplay of two different mechanisms, PRL or ovarian steroid hormones independently or in combination in a tissue-specific manner. Furthermore, we found that ovarian steroid hormones selectively suppress the expression of PRLR isoforms in mouse fallopian tubes. These findings may contribute to our understanding of the mechanisms controlling PRLR isoform expression in the fallopian tube (in addition to ovary and uterus), with implications for female reproduction.
Unité(s) : U845 (VG)
 
  Rational design of competitive prolactin/growth hormone receptor antagonists
TALLET E, ROUET V, JOMAIN JB, KELLY PA, BERNICHTEIN S, GOFFIN V
2008 - J. Mammary Gland Biol. Neoplasi. 13(1):105-117
There is increasing evidence that prolactin (PRL) and growth hormone (GH) act as growth-promoters of breast tumors. Recent arguments have accumulated to suggest that when they are locally-produced within the mammary tissue, these hormones, acting by an autocrine-paracrine mechanism may have enhanced, or even specific functions compared to endocrine PRL and GH. Classical drugs blocking pituitary hormone production (dopamine and somatostatin analogs) are ineffective on extrapituitary expression of PRL/GH genes, therefore the undesirable effects of these locally-produced hormones remain a target of interest for alternative strategies. This has encouraged the development of competitive PRL and/or GH receptor antagonists, which involve engineered variants of natural receptor ligands (PRL or GH) aimed at blocking receptor activation rather than hormone production in peripheral tissues. This article overviews the rational design of this new class of molecules, their specific molecular features (receptor specificity, biological properties, etc.) and whenever available, the data that have been obtained in cell or animal models of breast cancer.
Unité(s) : U845 (VG)
 
  Polyubiquitination of prolactin receptor stimulates its internalization, postinternalization sorting, and degradation via the lysosomal pathway
VARGHESE B, BARRIERE H, CARBONE CJ, BANERJEE A, SWAMINATHAN G, PLOTNIKOV A, XU P, PENG J, GOFFIN V, LUKACS GL, FUCHS SY
2008 - Mol. Cell. Biol. 28(17):5275-5287
The ubiquitination of the receptor that mediates signaling induced by the polypeptide pituitary hormone prolactin (PRL) has been shown to lead to the degradation of this receptor and to the ensuing negative regulation of cellular responses to PRL. However, the mechanisms of PRL receptor (PRLr) proteolysis remain largely to be determined. Here we provide evidence that PRLr is internalized and primarily degraded via the lysosomal pathway. Ubiquitination of PRLr is essential for the rapid internalization of PRLr, which proceeds through a pathway dependent on clathrin and the assembly polypeptide 2 (AP2) adaptor complexes. Recruitment of AP2 to PRLr is stimulated by PRLr ubiquitination, which also is required for the targeting of already internalized PRLr to the lysosomal compartment. While mass spectrometry analysis revealed that both monoubiquitination and polyubiquitination (via both K48- and K63-linked chains) occur on PRLr, the results of experiments using forced expression of ubiquitin mutants indicate that PRLr polyubiquitination via K63-linked chains is important for efficient interaction of PRLr with AP2 as well as for efficient internalization, postinternalization sorting, and proteolytic turnover of PRLr. We discuss how specific ubiquitination may regulate early and late stages of endocytosis of PRLr and of related receptors to contribute to the negative regulation of the magnitude and duration of downstream signaling.
Unité(s) : U845 (VG)
 
  The growth hormone receptor in growth
BOUGNERES P, GOFFIN V
2007 - Endocrinol. Metab. Clin. North. Am. 36(1):1-16
The growth hormone receptor (GHR) is a major effector of Human growth. Functional variants of the GHR include very rare loss-of-function mutations (pathology) and very common polymorphisms (physiology). Recent experimental data have clarified the mechanisms through which mutations of the GHR or Stat5 lead to growth hormone insensitivity and major monogenic growth defects. Recent pharmacogenetic studies support that the response to growth-promoting administration of growth hormone is influenced by exon 3 polymorphism of the GHR.
Unité(s) : U845 (VG)
 
  Elevated Serum Bioactive Prolactin Concentrations in Patients with Systemic Lupus Erythematosus Are Associated with Disease Activity as Disclosed by Homologous Receptor Bioassays
CARDENAS-MONDRAGON G, ULLOA-AGUIRRE A, ISORDIA-SALAS I, GOFFIN V, LEANOS-MIRANDA A
2007 - J. Rheumatol. 34(7):1514-1521
OBJECTIVE: To assess the bioactivity of circulating prolactin (PRL) in serum samples from patients with systemic lupus erythematosus (SLE) using 2 novel homologous in vitro bioassays, and to correlate PRL bioactivity with lupus activity. METHODS: Serum samples from 98 SLE patients with and without disease activity were tested for immunoreactive and bioactive concentrations of PRL. RESULTS: Patients with active disease exhibited higher bioactive serum PRL levels in homologous bioassays (p
Unité(s) : U845 (VG)
 
  Prolactin stimulates prostate cell proliferation by increasing endoplasmic reticulum content due to SERCA 2b over-expression
CREPIN A, BIDAUX G, VANDEN-ABEELE F, DEWAILLY E, GOFFIN V, PREVARSKAYA N, SLOMIANNY C
2007 - Biochem. J. 401(1):49-55
Prolactin (PRL) has been shown to be involved in the differentiation and proliferation of numerous tissues, including the prostate gland. Moreover, variations in [Ca2+]ER (calcium concentration within the endoplasmic reticulum) may play a role in cell growth. However, few studies have focused on the regulation of calcium homoeostasis by prolactin. The present study evaluates the regulation of calcium pools as well as the possible role of [Ca2+]ER variations as a signal for growth modulation by PRL. We show that PRL stimulates the proliferation of normal SV40 immortalized epithelial prostate (PNT1A) cells with a maximum effect at a dose of 100 ng/ml. We also show that 100 ng/ml PRL increases the [Ca2+]ER when measured either by indirect quantification with Fura-2AM after application of 1 mM thapsigargin or by direct quantification with Mag-Fura-2AM within the endoplas-mic reticulum. Western blot analysis shows that the SERCA 2b (sarcoendoplasmic calcium ATPase 2b) is over-expressed in PNT1A cells treated with 100 ng/ml PRL for 24 h. A small inter-fering RNA SERCA 2a/b, used to down-regulate endogenous SERCA 2b expression, reduced both PNT1A cell proliferation and [Ca2+]ER. We thus identify [Ca2+]ER and SERCA 2b as protagonists in PRL-induced proliferation.
Unité(s) : U845 (VG)
 
  Autocrine Prolactin Promotes Prostate Cancer Cell Growth via Janus Kinase-2-Signal Transducer and Activator of Transcription-5a/b Signaling Pathway
DAGVADORJ A, COLLINS S, JOMAIN JB, ABDULGHANI J, KARRAS J, ZELLWEGER T, LI H, NURMI M, ALANEN K, MIRTTI T, VISAKORPI T, BUBENDORF L, GOFFIN V, NEVALAINEN MT
2007 - Endocrinology 148(7):3089-3101
The molecular mechanisms that promote progression of localized prostate cancer to hormone-refractory and disseminated disease are poorly understood. Prolactin (Prl) is a local growth factor produced in high-grade prostate cancer, and exogenously added Prl in tissue or explant cultures of normal and malignant prostate is a strong mitogen and survival factor for prostate epithelium. The key signaling proteins that mediate the biological effects of Prl in prostate cancer are Signal Transducer and Activator of Transcription (Stat)-5a/5b via activation of Janus kinase-2. Importantly, inhibition of Stat5a/b in prostate cancer cells induces apoptotic death. Using a specific Prl receptor antagonist (Delta1-9G129R-hPRL), we demonstrate here for the first time that autocrine Prl in androgen-independent human prostate cancer cells promotes cell viability via Stat5 signaling pathway. Furthermore, we examined a unique clinical material of human hormone refractory prostate cancers and metastases and show that autocrine Prl is expressed in 54% of hormone-refractory clinical human prostate cancers and 62% prostate cancer metastases. Finally, we demonstrate that autocrine Prl is expressed from both the proximal and distal promoters of the Prl gene in clinical human prostate cancers and in vivo and in vitro human prostate cancer models, independently of pituitary transcription factor-1 (Pit-1). Collectively, the data provide novel evidence for the concept that autocrine Prl signaling pathway is involved in growth of hormone-refractory and metastatic prostate cancer. The study also provides support for the use of Prl receptor antagonists or other therapeutic strategies to block the Prl-Janus kinase-2-Stat5 signaling pathway in advanced prostate cancer.
Unité(s) : U845 (VG)
 
  Structural and thermodynamic bases for the design of pure prolactin receptor antagonists: X-ray structure of Del1-9-G129R-hPRL
JOMAIN JB, TALLET E, BROUTIN I, HOOS S, VAN AGTHOVEN J, DUCRUIX A, KELLY PA, KRAGELUND BB, ENGLAND P, GOFFIN V
2007 - J. Biol. Chem. 282(45):33118-33131
Competitive antagonists of the human prolactin (hPRL) receptor are a novel class of molecules of potential therapeutic interest in the context of cancer. We recently developed the pure antagonist Del1-9-G129R-hPRL by deleting the nine N-terminal residues of G129R-hPRL, a first generation partial antagonist. We determined the crystallographic structure of Del1-9-G129R-hPRL, which revealed no major change compared with wild type hPRL, indicating that its pure antagonistic properties are intrinsically due to the mutations. To decipher the molecular bases of pure antagonism, we compared the biological, physicochemical, and structural properties of numerous hPRL variants harboring N-terminal or Gly(129) mutations, alone or combined. The pure versus partial antagonistic properties of the multiple hPRL variants could not be correlated to differences in their affinities toward the hPRL receptor, especially at site 2 as determined by surface plasmon resonance. On the contrary, residual agonism of the hPRL variants was found to be inversely correlated to their thermodynamic stability, which was altered by all the Gly(129) mutations but not by those involving the N terminus. We therefore propose that residual agonism can be abolished either by further disrupting hormone site 2-receptor contacts by N-terminal deletion, as in Del1-9-G129R-hPRL, or by stabilizing hPRL and constraining its intrinsic flexibility, as in G129V-hPRL.
Unité(s) : U845 (VG)
 
  Guidelines of the Pituitary Society for the diagnosis and management of prolactinomas
CASANUEVA FF, MOLITCH ME, SCHLECHTE JA, ABS R, BONERT V, BRONSTEIN MD, BRUE T, CAPPABIANCA P, COLAO A, FAHLBUSCH R, FIDELEFF H, HADANI M, KELLY PA, KLEINBERG D, LAWS E, MAREK J, SCANLON M, SOBRINHO LG, WASS JAH, GIUSTINA A
2006 - Clin. Endocrinol. 65(2):265-273
In June 2005, an ad hoc Expert Committee formed by the Pituitary Society convened during the 9th International Pituitary Congress in San Diego, California. Members of this committee consisted of invited international experts in the field, and included endocrinologists and neurosurgeons with recognized expertise in the management of prolactinomas. Discussions were held that included all interested participants to the Congress and resulted in formulation of these guidelines, which represent the current recommendations on the diagnosis and management of prolactinomas based upon comprehensive analysis and synthesis of all available data.
Unité(s) : U808
 
  Prolactin modulates TRPV1 in female rat trigeminal sensory neurons
DIOGENES A, PATWARDHAN AM, JESKE NA, RUPAREL NB, GOFFIN V, AKOPIAN AN, HARGREAVES KM
2006 - J. Neurosci. 26(31):8126-8136
Sex dependency in pain perception is well documented and is thought to be attributable to the effect of reproductive hormones on nociceptive processing. In the present study, we evaluated whether estradiol alters gene transcription in the trigeminal ganglia (TG) of ovariectomized rats (OVX). These experiments demonstrated a dramatic (40-fold) upregulation of prolactin (PRL) expression in TG by 17-beta-estradiol (E2). PRL expression was restricted to TG neurons and was highly overlapped with transient potential receptor vanilloid type 1 (TRPV1) (approximately 90%) in TG. Additionally, PRL is released from neurons during stimulation. Both forms of PRL receptors (PRLRs), short and long, were also present in TG neurons. Moreover, expression of the long PRLRs was under control of estradiol. We next evaluated the novel hypothesis that PRL acts as a neuromodulator of sensory neurons. PRL pretreatment significantly enhanced capsaicin-evoked inward currents, calcium influx, and immunoreactive calcitonin gene-related peptide release from cultured TG neurons. This PRL modulation of capsaicin responses was abolished by withdrawal of E2 from TG cultures. Biochemical analysis demonstrated that PRL increased (>50%) phosphorylation levels of TRPV1 in TG. In a behavioral test, PRL pretreatment significantly potentiated capsaicin-evoked nocifensive behavior in female rats at proestrous and in OVX rats after E2 treatment. The in vivo potentiating effect of PRL on capsaicin responses was also dependent on E2. Collectively, these data demonstrate that PRL is a novel modulator of sensory neurons tightly regulated by E2. These findings are consistent with the hypothesis that PRL could contribute to the development of certain pain disorders, possibly including those modulated by estrogen.
Unité(s) : U808
 
  Human Macroprolactin Displays Low Biological Activity via Its Homologous Receptor in a New Sensitive Bioassay
GLEZER A, SOARES CR, VIEIRA JG, GIANNELLA-NETO D, RIBELA MT, GOFFIN V, BRONSTEIN MD
2006 - J. Clin. Endocrinol. Metabol. 91(3):1048-1055
Context: Macroprolactinemia is a frequent finding in hyperprolactinemic individuals, usually without clinical impact. Data on biological activity of macroprolactin (bbPRL) are controversial and mostly based on a heterologous rat Nb2 cell bioassay. Biological activity of bbPRL observed in vitro but not in vivo may be due to its high molecular weight, preventing its passage through capillary barrier. Alternatively, bbPRL bioactivity may differ depending on the prolactin (PRL) receptor (PRLR) species specificity. Objective: The objective of the study was to characterize the bioactivity of bbPRL in a homologous bioassay: Ba/F-3 cells stably expressing the human PRLR. Design/Setting/Patients: Chromatography-purified bbPRL from macroprolactinemic individuals (group I, n = 18) and monomeric PRL from hyperprolactinemic patients without macroprolactinemia (group II, n = 5) were tested in Nb2 and Ba/F-LLP bioassays. Both groups were followed up at the neuroendocrinology outpatients' clinic. Main Outcome Measure: Biological activity of bbPRL presented in the two bioassays was measured. Results: In group I, no patient had hypogonadism. Mean ratio bioactivity to immunoactivity of bbPRL in the Nb2 assay was 0.69. There was no dose-response in 15 of the 18 samples tested in Ba/F-LLP assay. In group II, three patients had galactorrhea and all five had hypogonadism. Mean ratio bioactivity to immunoactivity of monomeric PRL samples was 1.35 in Nb2 and 0.91 in Ba/F-LLP assay. Conclusion: Whereas both bioassays achieve similar results with respect to monomeric PRL activity, our results indicate that the activity displayed by bbPRL toward the rat receptor may be inappropriate because it is not observed in the human PRLR-mediated assay, consistent with the apparent absence of bioactivity in vivo.
Unité(s) : U808
 
  Drug Insight: prolactin-receptor antagonists, a novel approach to treatment of unresolved systemic and local hyperprolactinemia ?
GOFFIN V, TOURAINE P, CULLER MD, KELLY PA
2006 - Nat. Clin. Pract. Endocrinol. Metab. 2(10):571-581
Prolactin is a polypeptide hormone whose major biological actions are related to normal lactation and reproduction. Abnormally high prolactin levels, referred to as hyperprolactinemia, can result in various reproductive disorders. Currently, therapeutic management of hyperprolactinemia relies on dopamine agonists, since dopamine is the primary physiological suppressor of pituitary prolactin production. Epidemiologic studies have shown that prolactin levels in the high-normal range, as well as medications that interfere with dopamine action (e.g. certain antipsychotic drugs), might correlate with increased breast cancer risk. In addition to circulating prolactin, it is now well established that prolactin is also produced locally within various tissues, including breast and prostate. Increasing evidence, mainly from animal studies at present, suggests that excess locally produced prolactin may promote the growth of breast and prostate tumors via an autocrine or paracrine mechanism. These findings have renewed the interest in finding alternative strategies to suppress prolactin actions when dopamine agonists are ineffective. Our studies of the relationship between prolactin structure and function have resulted in the development of pure prolactin-receptor antagonists. These molecules prevent endogenous prolactin from exerting its actions via a competitive mechanism for receptor binding. In this review, we discuss the possible future therapeutic utility of this novel class of compounds.
Unité(s) : U808
 
  Application of new homologous in vitro bioassays for human lactogens to assess the actual bioactivity of human prolactin isoforms in hyperprolactinaemic patients
LEANOS-MIRANDA A, CARDENAS-MONDRAGON G, RIVERA-LEANOS R, ULLOA-AGUIRRE A, GOFFIN V
2006 - Clin. Endocrinol. 65(2):146-153
Background Prolactin (PRL) plays a central role in mammary gland development and lactation. Due to its molecular heterogeneity, measurement of PRL immunoreactivity does not necessarily reflect its intrinsic bioactivity. For many years the Nb2 rat lymphoma cell bioassay has been the only reference bioassay for human lactogens. This bioassay, however, does not always correlate with the clinical features found in some patients exhibiting normal or elevated immunoreactive serum PRL concentrations. Objectives (1) To determine the concentrations of bioactive PRL in serum samples from individuals with normoprolactinaemia or with different forms of hyperprolactinaemia using two recently described homologous in vitro bioassays (i.e. a transcriptional bioassay in HEK-293 cells and a proliferation assay in Ba/F3 cells); and (2) to compare these results with those generated by the classical Nb2 cell bioassay. Design Cross-sectional study. Setting An institutional biomedical research laboratory. Participants Ten patients with symptomatic hyperprolactinaemia due to prolactinoma, 11 patients with asymptomatic hyperprolactinaemia and macroprolactinaemia, and nine normal women. Main outcome measures Measurement of immunoreactive and bioactive concentrations of serum PRL. Results Samples from normal women and patients with tumoral hyperprolactinaemia due to prolactinoma exhibited similar within-group concentration values of bioactive and immunoreactive serum PRL when tested by the three bioassays and the immunoradiometric assay employed. By contrast, measurement of bioactive PRL in samples from patients with macroprolactinaemia revealed that macroprolactin was poorly active in the homologous receptor bioassays, while it was more active in the Nb2 bioassay. Conclusions The reduced bioactivity of PRL in patients with macroprolactinaemia may further explain the absence of clinical features of hyperprolactinaemia in these individuals. In addition, our findings indicate that species-specificity and sensitivity of the bioassays are determinant factors in the measurement of the intrinsic biological activity of circulating PRL.
Unité(s) : U808
 
  Local over-expression of prolactin in differentiating mouse mammary gland induces functional defects and benign lesions, but no carcinoma
MANHS C, KAYSER C, BERTHEAU P, KELDER B, KOPCHICK JJ, KELLY PA, TOURAINE P, GOFFIN V
2006 - J. Endocrinol. 190(2):271-285
Experimental, clinical, and epidemiological data support the growth-promoting role of endocrine prolactin (PRL) in mammary tumors. PRL is also produced by the breast, where it is now recognized to act as a growth/survival factor via autocrine/paracrine mechanisms. Recent transgenic (Tg) mouse models have revealed the pro-oncogenic effect of PRL over-expression in virgin mammary glands. To address the question whether PRL tumorigenicity was maintained on differentiated mammary glands, we generated mammary-specific Tg mice expressing human (h)PRL under the control of the milk whey acidic protein promoter, which directs autocrine hPRL over-expression in late gestation throughout lactation. Minimal levels of transgene expression were detected in the mammary glands of virgin animals, which at best induced partial ductal branching and lobulo-alveolar structures in older nulliparous females. As expected, expression of mammary hPRL dramatically increased at the end of first pregnancy, and from this point it never returned to baseline, although it peaked at each gestation/lactation cycle. Over-expression of hPRL that starts when the gland is already well into the differentiation process led to various morphological mammary alterations, including abnormally differentiated epithelium, atropy of the myoepithelial layer, dilated ducts, cysts, and lymphocytic infiltrates. These phenotypes tended to worsen with successive pregnancies, also reflecting cumulative damage of failure of involution. Although some older, multiparous females developed benign tumors (papillomas and metaplasias), none of the animals studied developed mammary carcinomas. In addition, we noticed that half of the Tg females exhibited lactation defects, leading to significantly increased pup mortality. This phenotype was due neither to failure of milk production nor to modification of its protein content, but rather it was correlated to lipid enrichment of the milk, which, in combination with profoundly altered morphology of the gland, led to impaired milk extrusion through the nipple. In summary, these data show that over-expression of autocrine hPRL in a differentiating mammary gland induces dramatic functional and morphological defects, but not carcinoma. This deserves further investigations on the emerging concept that autocrine PRL may have different effects on pathological development of the mammary gland depending on the differentiation state of the latter.
Unité(s) : U808
 
  A New Mechanism for Prolactin Processing into 16K PRL by Secreted Cathepsin D
PIWNICA D, FERNANDEZ I, BINART N, TOURAINE P, KELLY PA, GOFFIN V
2006 - Mol. Endocrinol. 20(12):3263-3278
Cathepsins are lysosomal enzymes that were shown to release the antiangiogenic fragments 16K prolactin (PRL), endostatin, and angiostatin by processing precursors at acidic pH in vitro. However, the physiological relevance of these findings is questionable because the neutral pH of physiological fluids is not compatible with the acidic conditions required for the proteolytic activity of these enzymes. Here we show that cathepsin D secreted from various tissues is able to process PRL into 16K PRL outside the cell. To specifically target extracellular proteolysis, we used tissues from PRL receptor-deficient mice, which are unable to internalize PRL. As assessed by the use of specific inhibitors of proton extruders, we show that the proteolytic activity of cathepsin D requires local acid secretion driven by Na(+)/H(+) exchangers and H(+)/ATPase. Although it is usually assumed that cathepsin-mediated generation of antiangiogenic peptides occurs in the moderately acidic pericellular milieu found in malignant tumors, we propose a new mechanism explaining the extracellular activity of this acidic protease under physiological pH. Our data support the concept that secreted lysosomal enzymes could be involved in the maintenance of angiogenesis dormancy via the generation of active antiangiogenic peptides in nonpathological contexts.
Unité(s) : U808, U809
 
  APRO4 negatively regulates Src tyrosine kinase activity in PC12 cells
RAHMANI Z
2006 - J. Cell Sci. 119(Pt 4):646-658
The Src nonreceptor tyrosine kinase plays an important role in multiple signalling pathways that regulate several cellular functions including proliferation, differentiation and transformation. The activity of Src is tightly regulated in vivo and can be modulated by interactions of its SH2 and SH3 domains with high-affinity ligands. APRO4 (anti-proliferative 4) belongs to a new antiproliferative gene family involved in the negative control of the cell cycle. This report shows that APRO4 associates with Src via its C-terminal proline-rich domain, and downregulates Src kinase activity. Moreover, overexpression of APRO4 leads to inhibition of neurite outgrowth and Ras/MAP kinase signalling in PC12 cells. Furthermore, the kinetics of endogenous Src inactivation correlates with an increase in endogenous APRO4 co-immunoprecipitation in FGF-stimulated PC12 cells. Finally, downregulation of endogenous APRO4 by expression of antisense RNA induces the activation of Src and spontaneous formation of neurites in PC12 cells. Therefore, by controlling the basal threshold of Src activity, APRO4 constitutes an important negative regulatory mechanism for Src-mediated signalling.
Unité(s) : U808
 
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