Publications by authors named "Florence Roucher-Boulez"

17 Publications

  • Page 1 of 1

Hypopituitarism in Patients with Blepharophimosis and FOXL2 Mutations.

Horm Res Paediatr 2020 26;93(1):30-39. Epub 2020 May 26.

Hospices Civils de Lyon, Hôpital Femme Mère Enfant, Service d'Endocrinologie Pédiatrique, Bron, France.

Background: FOXL2 is the gene involved in blepharophimosis, ptosis, and epicanthus inversus syndrome (BPES). There have been few single case reports of growth hormone deficiency (GHD) with this syndrome, and Foxl2 is known to be involved in pituitary development in mice. Our aim was to analyze the prevalence of FOXL2 gene alteration in a series of patients with congenital hypopituitarism and eyelid anomalies.

Methods: FOXL2 was analyzed in 10 patients with hypopituitarism (ranging from isolated GHD to complete pituitary hormone deficiency) and eyelid anomalies (typical BPES in 4 patients and milder anomalies in 6 patients). In patients with an FOXL2 mutation, we ruled out other possible molecular explanations by analyzing a panel of 20 genes known to be associated with hypopituitarism, and a candidate gene approach was used for patients without an FOXL2mutation.

Results: Three patients had an FOXL2mutation. All 3 had typical BPES. Their pituitary phenotype varied from GHD to complete pituitary hormone deficiency and their pituitary morphology ranged from normal to an interrupted pituitary stalk. No mutations were found in genes previously associated with hypopituitarism.

Conclusion: Our study shows that some patients with BPES have hypopituitarism with no molecular explanation other than FOXL2 mutation. This points toward an involvement of FOXL2 in human pituitary development.
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http://dx.doi.org/10.1159/000507249DOI Listing
May 2020

Age- and Sex-Specific TSH Upper-Limit Reference Intervals in the General French Population: There Is a Need to Adjust Our Actual Practices.

J Clin Med 2020 Mar 14;9(3). Epub 2020 Mar 14.

Hospices Civils de Lyon, Fédération d'Endocrinologie, Groupement Hospitalier Est, F-69677 Bron cedex, France.

It is well known that thyroid dysfunction increases with age. This study is aimed to determine reference intervals, in males and females, suitable for thyroid disease exploration during adult life using routinely collected serum thyrotropin (TSH) data in a tertiary center from 2007 to 2018. Over 11 years, 295,775 TSH levels were measured in a single lab. Among the 156,025 TSH results available for analysis, 90,538 values were from female subjects, 82,019 were from patients aged >60 years and 26,825 were from patients aged >80 years. By using an indirect approach, we determined reference values of TSH adapted to age and sex, and we then evaluated the proportion of patients who would have been reclassified with these reference values. The median TSH ranged from 1.2-1.4 mUI/L during the study period. The upper limit of reference range of TSH increased with age; in females the median to 97.5th percentile values increased continuously from the age of 30 years to the oldest age group. Using new calculated reference values in patients with TSH above the conventional upper-limit reference value (4 mUI/L), the proportion of results reclassified as within the reference interval among patients aged >60 years ranged, according to age group, from 50.5% to 65.1% of females and from 33.0% to 37.7% of males. The use of TSH age-specific and sex-specific upper-limit reference values led to the reclassification of a great number of samples, notably among women. This suggests that age-specific TSH upper-limit reference intervals in daily practice should be used in order to avoid misclassification.
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http://dx.doi.org/10.3390/jcm9030792DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7141356PMC
March 2020

Reversion SAMD9 Mutations Modifying Phenotypic Expression of MIRAGE Syndrome and Allowing Inheritance in a Usually Disorder.

Front Endocrinol (Lausanne) 2019 11;10:625. Epub 2019 Sep 11.

Laboratoire de Biochimie et Biologie Moléculaire Grand Est, UM Pathologies Endocriniennes Rénales Musculaires et Mucoviscidose, Groupement Hospitalier Est, Hospices Civils de Lyon, Bron, France.

MIRAGE (Myelodysplasia, Infection, Restriction of growth, Adrenal hypoplasia, Genital phenotypes, Enteropathy) syndrome is a severe multisystem disorder with high mortality. It is caused by a heterozygous gain of function mutation in the growth repressor gene . The increasing number of reported cases displays a spectrum of phenotypes that may be explained by an adaptation mechanism, with appearance of a somatic second hit mutation with revertant effects. To determine the genetic basis of the MIRAGE syndrome rapidly corrected in a living and healthy 46,XY patient. A 46,XY patient born with growth restriction and disorders of sex development had thrombocytopenia and necrotizing enterocolitis during the neonatal period suggestive of the syndrome. Faced with the rapid improvement of the patient's phenotype, an adaptation mechanism was sought by repeating genetic analysis at different ages; her parents also underwent genetic analysis. The previously described p.(Thr778Ile) mutation was identified and surprisingly transmitted by the asymptomatic mother in this usually syndrome. To explain the rapid improvement of the patient's phenotype and absence of symptoms in the mother, an adaptation mechanism was sought. For the mother, a non-sense mutation was found (p.(Arg221)) in , and most likely appeared . It was not transmitted to her child. The child harbored a different non-sense mutation (p.(Arg285)) that most likely appeared near day 20. We show that pathogenic variants can be inherited from a healthy parent as the adaptation mechanism may arise early in life and mask symptoms. Presence of revertant mosaicism mutations could explain "incomplete penetrance" in other disease. For a better management and outcomes in patients, appearance of this natural gene therapy should be sought by repeating genetic analysis.
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http://dx.doi.org/10.3389/fendo.2019.00625DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6749008PMC
September 2019

Hormonal and spatial control of SUMOylation in the human and mouse adrenal cortex.

FASEB J 2019 09 17;33(9):10218-10230. Epub 2019 Jun 17.

Génétique Reproduction and Dévelopement (GReD), Centre National de la Recherche Scientifique (CNRS), INSERM, Université Clermont-Auvergne, Clermont-Ferrand, France.

SUMOylation is a highly conserved and dynamic post-translational mechanism primarily affecting nuclear programs for adapting organisms to stressful challenges. Alteration of SUMOylation cycles leads to severe developmental and homeostatic defects and malignancy, but signals coordinating SUMOylation are still unidentified. The adrenal cortex is a zonated endocrine gland that controls body homeostasis and stress response. Here, we show that in human and in mouse adrenals, SUMOylation follows a decreasing centripetal gradient that mirrors cortical differentiation flow and delimits highly and weakly SUMOylated steroidogenic compartments, overlapping glomerulosa, and fasciculata zones. Activation of PKA signaling by acute hormonal treatment, mouse genetic engineering, or in Carney complex results in repression of small ubiquitin-like modifier (SUMO) conjugation in the inner cortex by coordinating expression of SUMO pathway inducers and repressors. Conversely, genetic activation of canonical wingless-related integration site signaling maintains high SUMOylation potential in the outer neoplastic cortex. Thus, SUMOylation is tightly regulated by signaling pathways that orchestrate adrenal zonation and diseases.-Dumontet, T., Sahut-Barnola, I., Dufour, D., Lefrançois-Martinez, A.-M., Berthon, A., Montanier, N., Ragazzon, B., Djari, C., Pointud, J.-C., Roucher-Boulez, F., Batisse-Lignier, M., Tauveron, I., Bertherat, J., Val, P., Martinez, A. Hormonal and spatial control of SUMOylation in the human and mouse adrenal cortex.
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http://dx.doi.org/10.1096/fj.201900557RDOI Listing
September 2019

Isolated hypoaldosteronism as first sign of X-linked adrenal hypoplasia congenita caused by a novel mutation in NR0B1/DAX-1 gene: a case report.

BMC Med Genet 2019 06 4;20(1):98. Epub 2019 Jun 4.

Laboratoire de Biochimie et Biologie Moléculaire Grand Est, UM Pathologies Endocriniennes Rénales Musculaires et Mucoviscidose, Groupement Hospitalier Est, Hospices Civils de Lyon, Univ Lyon, Université Claude Bernard Lyon 1, Lyon, France.

Background: X-linked Adrenal Hypoplasia Congenita (AHC) is a rare cause of primary adrenal insufficiency due to mutations in the NR0B1 gene, causing a loss of function of the nuclear receptor protein DAX-1. Adrenal insufficiency usually appears in the first 2 months of life, but can sometimes emerge during childhood. Hypogonadotropic Hypogonadism is often associated later in life and patients may develop azoospermia. We describe an unusual onset of AHC started with isolated hypoaldosteronism as first and only sign of the disease.

Case Presentation: A 18-days-old newborn presented with failure to thrive and feeding difficulties. Blood tests showed severe hyponatremia, hyperkalemia and hypochloremia. Renin was found over the measurable range and aldosterone was low whereas cortisol level was normal with a slightly increased ACTH. In the suspicion of Primary Hypoaldosteronism, correction of plasmatic electrolytes and replacement therapy with Fludrocortisone were promptly started. The subsequent evidence of low plasmatic and urinary cortisol and increased ACTH required the start of Hydrocortisone replacement therapy and it defined a clinical picture of adrenal insufficiency. Genetic analysis demonstrated a novel mutation in the DAX-1 gene leading to the diagnosis of AHC.

Conclusions: AHC onset may involve the aldosterone production itself, miming an isolated defect of aldosterone synthesis. NR0B1/DAX-1 mutations should be considered in male infants presenting with isolated hypoaldosteronism as first sign of adrenal insufficiency.
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http://dx.doi.org/10.1186/s12881-019-0834-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6549270PMC
June 2019

Letter to the Editor: "Characterization of the CYP11A1 Nonsynonymous Variant p.E314K in Children Presenting With Adrenal Insufficiency".

J Clin Endocrinol Metab 2019 05;104(5):1413-1414

Laboratoire de Biochimie et Biologie Moléculaire Grand Est, UM Pathologies Endocriniennes Rénales Musculaires et Mucoviscidose, Groupement Hospitalier Est, Hospices Civils de Lyon, Bron, France.

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http://dx.doi.org/10.1210/jc.2018-02415DOI Listing
May 2019

Aberrant Splicing Is the Pathogenicity Mechanism of the p.Glu314Lys Variant in Gene.

Front Endocrinol (Lausanne) 2018 5;9:491. Epub 2018 Sep 5.

Laboratoire de Biochimie et Biologie Moléculaire Grand Est, UM Pathologies Endocriniennes Rénales Musculaires et Mucoviscidose, Groupement Hospitalier Est, Hospices Civils de Lyon, Bron, France.

The cholesterol side chain cleavage enzyme (CYP11A1) catalyzes the conversion of cholesterol to pregnenolone, the first rate-limiting step of steroidogenesis. mutations are associated with primary adrenal insufficiency (PAI) as well as disorders of sex development (DSD) in 46,XY patients. To define the pathogenicity mechanism for the p.Glu314Lys variant, previously reported, and found in four additional patients with CYP11A1 deficiency. DNA of four patients presenting with delayed PAI and/or 46,XY DSD were studied by Sanger or Massively Parallel sequencing. Three mutations were characterized and , and one by mRNA analysis on testicular tissue. All patients were compound heterozygous for the previously described p.Glu314Lys variant. studies predicted this mutation as benign with no effect on splicing but mRNA analysis found that it led to incomplete exon 5 skipping. This mechanism was confirmed by minigene experiment. The protein carrying this mutation without exon skipping should conserve almost normal activity, according to studies. Two other mutations found in , the p.Arg120Gln and p.Arg465Trp, had similar activity compared to negative control, consistent with the studies. We provide biological proof that the p. Glu314Lys variant is pathogenic due to its impact on splicing and seems responsible for the moderate phenotype of the four patients reported herein. The present study highlights the importance of considering the potential effect of a missense variant on splicing when it is not predicted to be disease causing.
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http://dx.doi.org/10.3389/fendo.2018.00491DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6134065PMC
September 2018

Adrenocortical development: Lessons from mouse models.

Ann Endocrinol (Paris) 2018 Jun 16;79(3):95-97. Epub 2018 Apr 16.

GReD, CNRS, Inserm, université Clermont-Auvergne, 63001 Clermont-Ferrand, France. Electronic address:

The adrenocortical gland undergoes structural and functional remodelling in the fetal and postnatal periods. After birth, the fetal zone of the gland undergoes rapid involution in favor of the definitive cortex, which reaches maturity with the emergence of the zona reticularis(zR) at the adrenarche. The mechanisms underlying the adrenarche, the process leading to pre-puberty elevation of plasma androgens in higher primates, remain unknown, largely due to lack of any experimental model. By following up fetal and definitive cortex cell lines in mice, we showed that activation of protein kinase A (PKA) signaling mainly impacts the adult cortex by stimulating centripetal regeneration, with differentiation and then conversion of the zona fasciculata into a functional zR. Animals developed Cushing syndrome associated with primary hyperaldosteronism, suggesting possible coexistence of these hypersecretions in certain patients. Remarkably, all of these traits were sex-dependent: testicular androgens promoted WNT signaling antagonism on PKA, slowing cortical renewal and delaying onset of Cushing syndrome and the establishment of the zR in male mice, this being corrected by orchidectomy. In conclusion, zR derives from centripetal conversion of the zona fasciculata under cellular renewal induced by PKA signaling, determining the size of the adult cortex. Finally, we demonstrated that this PKA-dependent mobilization of cortical progenitors is sexually dimorphic and could, if confirmed in humans, account for female preponderance in adrenocortical pathologies.
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http://dx.doi.org/10.1016/j.ando.2018.03.014DOI Listing
June 2018

News about the genetics of congenital primary adrenal insufficiency.

Ann Endocrinol (Paris) 2018 Jun 13;79(3):174-181. Epub 2018 Apr 13.

Laboratoire de biochimie et biologie moléculaire Grand Est, UM pathologies endocriniennes rénales musculaires et mucoviscidose, groupement hospitalier Est, hospices civils de Lyon, 69677 Bron, France; Université de Lyon, université Claude-Bernard Lyon 1, 69008 Lyon, France.

Primary adrenal insufficiency (PAI) is characterized by impaired production of steroid hormones due to an adrenal cortex defect. This condition incurs a risk of acute insufficiency which may be life-threatening. Today, 80% of pediatric forms of PAI have a genetic origin but 5% have no clear genetic support. Recently discovered mutations in genes relating to oxidative stress have opened the way to research on genes unrelated to the adrenal gland. Identification of causal mutations in a gene responsible for PAI allows genetic counseling, guidance of follow-up and prevention of complications. This is particularly true for stress oxidative anomalies, as extra-adrenal manifestations may occur due to the sensitivity to oxidative stress of other organs such as the heart, thyroid, liver, kidney and pancreas.
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http://dx.doi.org/10.1016/j.ando.2018.03.016DOI Listing
June 2018

PKA signaling drives reticularis differentiation and sexually dimorphic adrenal cortex renewal.

JCI Insight 2018 01 25;3(2). Epub 2018 Jan 25.

GReD, Université Clermont Auvergne, CNRS, INSERM, Clermont-Ferrand, France.

The adrenal cortex undergoes remodeling during fetal and postnatal life. How zona reticularis emerges in the postnatal gland to support adrenarche, a process whereby higher primates increase prepubertal androgen secretion, is unknown. Using cell-fate mapping and gene deletion studies in mice, we show that activation of PKA has no effect on the fetal cortex, while it accelerates regeneration of the adult cortex, triggers zona fasciculata differentiation that is subsequently converted into a functional reticularis-like zone, and drives hypersecretion syndromes. Remarkably, PKA effects are influenced by sex. Indeed, testicular androgens increase WNT signaling that antagonizes PKA, leading to slower adrenocortical cell turnover and delayed phenotype whereas gonadectomy sensitizes males to hypercorticism and reticularis-like formation. Thus, reticularis results from ultimate centripetal conversion of adult cortex under the combined effects of PKA and cell turnover that dictate organ size. We show that PKA-induced progenitor recruitment is sexually dimorphic and may provide a paradigm for overrepresentation of women in adrenal diseases.
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http://dx.doi.org/10.1172/jci.insight.98394DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5821213PMC
January 2018

Triple-A syndrome: a wide spectrum of adrenal dysfunction.

Eur J Endocrinol 2018 Mar 13;178(3):199-207. Epub 2017 Dec 13.

Univ LyonUniversité Claude Bernard Lyon 1, Lyon, France.

Objective: Triple-A or Allgrove syndrome is an autosomal recessive disorder due to mutations in the gene, which encodes a nucleoporin named ALADIN. It is characterized by a classical clinical triad: alacrima, achalasia and adrenal insufficiency, the canonic symptoms that are associated with progressive peripheral neuropathy. Only a few cohorts have been reported. The objective of the present study was to characterize the various spectra of adrenal function in Triple-A patients.

Methods: A retrospective clinical and biological monitoring of 14 patients (10 families) was done in a single multidisciplinary French center. All had gene sequenced and adrenal function evaluation.

Results: Nine different mutations were found, including one new mutation: c.755G>C, p.(Trp252Ser). Regarding adrenal function, defects of the zona fasciculata and reticularis were demonstrated by increased basal ACTH levels and low DHEAS levels in all cases regardless of the degree of glucocorticoid deficiency. In contrast, mineralocorticoid function was always conserved: i.e., normal plasma renin level associated with normal aldosterone level. The main prognostic feature was exacerbation of neuropathy and cognitive disorders.

Conclusions: These data suggest that, in Triple-A patients, adrenal function can be deficient, insufficient or compensated. In our cohort after the first decade of life, there does not appear to be any degradation of adrenal function over time. However, patients with compensated adrenal function should be informed and educated to manage a glucocorticoid replacement therapy in case of stressful conditions, with no need for systematic long-term treatment.
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http://dx.doi.org/10.1530/EJE-17-0642DOI Listing
March 2018

Successful IVF pregnancy despite inadequate ovarian steroidogenesis due to congenital lipoid adrenal hyperplasia (CLAH): a case report.

Hum Reprod 2016 11 3;31(11):2609-2612. Epub 2016 Oct 3.

Aix-Marseille Université, CNRS, CRN2M UMR 7286, 13344 cedex 15, Marseille, France

Steroidogenic acute regulatory protein (StAR) mutations are the most frequent aetiologies of congenital lipoid adrenal hyperplasia (CLAH). Phenotypes may vary, and puberty may be absent in affected individuals. To date, only two pregnancies have been described in 46,XX CLAH patients with StAR mutations; these patients exhibited ovarian steroidogenesis along with spontaneous puberty and menarche and normal menses. The patient described here presented with CLAH caused by the homozygous (unreported, 1 bp) deletion c.719del in the StAR gene, which was diagnosed after acute adrenal insufficiency when the patient was 10 days old. The patient did not undergo spontaneous puberty, so puberty was induced by HRT when the patient was 13 years old. At the age of 25 years, the patient was referred to our reproductive unit because she desired to conceive. An initial cycle of clomiphene, stimulation produced follicular growth with two mature follicles measuring 18 and 15 mm, respectively, but the plasma oestradiol levels remained low (18 pg/ml) and the endometrium was thin (3 mm). Pregnancy was finally achieved after ovarian stimulation, IVF and transfer of frozen-thawed embryos after endometrial preparation with HRT. A normal female child was delivered following a 40 weeks' uneventful pregnancy. We therefore report the first IVF pregnancy achieved in a 46,XX CLAH patient homozygous for a StAR mutation, with inadequate ovarian steroidogenesis and no spontaneous puberty.
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http://dx.doi.org/10.1093/humrep/dew239DOI Listing
November 2016

P450 Oxidoreductase Deficiency: Loss of Activity Caused by Protein Instability From a Novel L374H Mutation.

J Clin Endocrinol Metab 2016 12 7;101(12):4789-4798. Epub 2016 Sep 7.

Pediatric Endocrinology, Diabetology, and Metabolism (S.P., C.E.F., A.V.P.), University Children's Hospital, and Department of Clinical Research, University of Bern, CH-3010 Bern, Switzerland; Service d'Endocrinologie Moléculaire et Maladies Rares (F.R.-B., D.M., Y.M.), Centre de Biologie et Pathologie Est, Hospices Civils de Lyon, Université Lyon 1, 69002 Bron-Lyon, France; and Endocrinologie Pédiatrique Département de Pédiatrie Médicale (A.L.-R.), Hôpital de la Mère et de l'Enfant, 87042 Limoges, France.

Context: P450 oxidoreductase (POR) is required for the activities of steroid-metabolizing cytochrome P450 enzymes in the endoplasmic reticulum. POR deficiency (PORD) is a form of congenital adrenal hyperplasia. Objective and Aim: Enzymatic and structural analysis of a novel L374H POR mutation from a patient with 46,XX disorder of sexual development. Design, Setting, Patient, and Intervention: The patient was a 46,XX girl with nonconsanguineous Turkish parents. She had virilized external genitalia at birth, a uterus and ovaries, and no sign of Antley-Bixler syndrome. The initial diagnosis was CYP21A2 deficiency with no mutations in CYP21A2, but POR mutations were found. Functional testing was done after producing recombinant POR proteins for analyzing enzymatic and structural properties.

Main Outcome: Novel mutations were causing severe loss of POR activities for metabolism of steroids and small molecules.

Results: The L374H mutation reduced activities by 80% in cytochrome c, 97% in thiazolyl blue tetrazolium bromide, and 86% in ferricyanide reduction assays. The catalytic efficiency of the 17 α-hydroxylation of progesterone and the 17,20-lyase reaction of 17-OH pregnenolone was decreased by 87 and 90%, respectively; 21-hydroxylation of progesterone was decreased by 96%, and androstenedione aromatization was decreased by 90%. Analysis of the mutant structure by molecular dynamics simulations revealed structural instability. Flavin release and fast proteolysis assays showed that the L374H mutant is less stable than wild-type POR, confirming the bioinformatics prediction.

Conclusions: This is the first report of a mutation causing PORD by affecting protein stability that causes severe reduction in POR activities. Detailed characterization of individual mutations in POR is required for understanding novel molecular mechanisms causing PORD.
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http://dx.doi.org/10.1210/jc.2016-1928DOI Listing
December 2016

NNT mutations: a cause of primary adrenal insufficiency, oxidative stress and extra-adrenal defects.

Eur J Endocrinol 2016 Jul 29;175(1):73-84. Epub 2016 Apr 29.

Molecular Endocrinology and Rare DiseasesLyon University Hospital, Bron, France Claude Bernard Lyon 1 UniversityLyon, France.

Objective: Nicotinamide nucleotide transhydrogenase (NNT), one of the several genes recently discovered in familial glucocorticoid deficiencies (FGD), is involved in reactive oxygen species detoxification, suggesting that extra-adrenal manifestations may occur, due to the sensitivity to oxidative stress of other organs rich in mitochondria. Here, we sought to identify NNT mutations in a large cohort of patients with primary congenital adrenal insufficiency without molecular etiology and evaluate the degree of adrenal insufficiency and onset of extra-adrenal damages.

Methods: Sanger or massive parallel sequencing of NNT and patient monitoring.

Results: Homozygous or compound heterozygous NNT mutations occurred frequently (26%, 13 unrelated families, 18 patients) in our cohort. Seven new mutations were identified: p.Met337Val, p.Ala863Glu, c.3G>A (p.Met1?), p.Arg129*, p.Arg379*, p.Val665Profs*29 and p.Ala704Serfs*19. The most frequent mutation, p.Arg129*, was found recurrently in patients from Algeria. Most patients were diagnosed belatedly (8-18 months) after presenting severe hypoglycemia; others experiencing stress conditions were diagnosed earlier. Five patients also had mineralocorticoid deficiency at onset. One patient had congenital hypothyroidism and two cryptorchidism. In follow-up, we noticed gonadotropic and genitalia impairments (precocious puberty, testicular inclusions, interstitial Leydig cell adenoma, azoospermia), hypothyroidism and hypertrophic cardiomyopathy. Intrafamilial phenotype heterogeneity was also observed.

Conclusions: NNT should be sequenced, not only in FGD, but also in all primary adrenal insufficiencies for which the most frequent etiologies have been ruled out. As NNT is involved in oxidative stress, careful follow-up is needed to evaluate mineralocorticoid biosynthesis extent, and gonadal, heart and thyroid function.
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http://dx.doi.org/10.1530/EJE-16-0056DOI Listing
July 2016

A splicing mutation in the DMD gene detected by next-generation sequencing and confirmed by mRNA and protein analysis.

Clin Chim Acta 2015 Aug 3;448:146-9. Epub 2015 Jul 3.

Laboratoire d'Endocrinologie Moléculaire et Maladies Rares, Hospices Civils de Lyon, Lyon, France.

Background: Dystrophinopathies, either the severe Duchenne Muscular Dystrophy (DMD) or the milder Becker Muscular Dystrophy (BMD), are X-linked recessive disorders caused by mutations in the DMD gene. DMD is one of the longest human genes. Large deletions or duplications account for 60-80% of the mutations. Remaining anomalies consist in point mutations or small rearrangements. Routinely, the molecular diagnosis is done by a Multiplex Ligation-dependent Probe Amplification (MLPA) or array Comparative Genome Hybridization (aCGH), followed, if negative, by Sanger sequencing of all exons.

Methods: In this study, massive parallel sequencing (MPS) or next generation sequencing (NGS) was used to make a rapid and costless molecular diagnosis in a young boy suspected of DMD.

Results: A small deletion: NM_004006.2:c.2803+5_2803+8del was identified. The diagnosis was performed in one single manipulation and within a week. The consequence of this intronic mutation is a skipping of exon 21 confirmed by mRNA and protein analysis.

Conclusions: NGS appears to be an efficient new strategy in DMD molecular diagnosis. It highlights the major evolution of the diagnostic strategy towards high throughput technologies, where bioinformatics analysis becomes the real challenge for variations detection. This is the first study reporting in vivo impact of this intronic mutation.
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http://dx.doi.org/10.1016/j.cca.2015.07.002DOI Listing
August 2015

New NOBOX mutations identified in a large cohort of women with primary ovarian insufficiency decrease KIT-L expression.

J Clin Endocrinol Metab 2015 Mar 16;100(3):994-1001. Epub 2014 Dec 16.

Inserm U693 (J.B., J.B., A.G.M., J.Y., N.B.), Le Kremlin-Bicêtre, F-94276, France; Université Paris-Sud (J.B., J.Y., N.B.), Faculté de Médecine Paris-Sud, Le Kremlin-Bicêtre, F-94276, France; Service d'Hormonologie, d'Endocrinologie Moléculaire et Des Maladies Rares (F.R.-B.), Centre De Biologie et Pathologie Est, Université Lyon 1, 69677 Bron, France; Unité de Gynécologie Endocrinienne (A.G.), Université Paris-Descartes, l'Assistance Publique-Hôpitaux de Paris, Hôpitaux Universitaires Paris Centre, 75014 Paris, France; l'Assistance Publique-Hôpitaux de Paris (H.B-G., J.Y., N.B.), Hôpital de Bicêtre, Service d'Endocrinologie et des Maladies de la Reproduction, Le Kremlin-Bicêtre, F-94276, France; Centre d'Aide Médicale à la Procréation (H.B-G.), CHI 94000 Créteil, France; Service de Biochimie et Génétique Moléculaire (K.A., C.B., C.D.), Hôpital Cochin, l'Assistance Publique-Hôpitaux de Paris Université Paris-Descartes, 75006 Paris, France; l'Assistance Publique-Hôpitaux de Paris (J.B., A.G.M.), Service de Génétique Moléculaire, Pharmacogénétique et Hormonologie, Le Kremlin-Bicêtre, F-94276, France; and Service d'Endocrinologie-Diabète-Nutrition (A.-C.H., B.D.), Centre Hospitalier Universitaire de Reims-Hôpital Robert-Debré, 51092 Reims, France.

Context: Primary ovarian insufficiency (POI) is a major cause of anovulation and infertility in women. This disease affects 1% of women before 40 years, and several genetic causes have been reported.

Objective: The aim of the study was to evaluate the prevalence of NOBOX mutations in a new large cohort of women with POI and to characterize these variants and identify a NOBOX novel target gene.

Patients And Methods: A total of 213 unrelated patients with POI were screened for NOBOX mutations, and luciferase reporter assays were performed for the mutations identified.

Results: We reported 3 novel and 2 recurrent heterozygous missense NOBOX rare variants found in 12 patients but not in 724 alleles from ethnic-matched individual women with occurrence of menopause at a normal age. Their functional impact had been tested on the classic growth differentiation factor-9 (GDF9) promoter and on KIT-L, a new NOBOX target gene. The p.Gly91Thr, p.Gly111Arg, p.Arg117Trp, p.Lys371Thr, and p.Pro619Leu mutations were deleterious for protein function.

Conclusions: In our series, 5.6% of the patients with POI displayed heterozygous NOBOX mutations. We demonstrate that KIT-L could be now a direct NOBOX target. These findings replicate the high prevalence of the association between the NOBOX rare variants and POI.
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http://dx.doi.org/10.1210/jc.2014-2761DOI Listing
March 2015