FGFR1 and PROKR2 rare variants found in patients with combined pituitary hormone deficiencies.

The genetic aetiology of congenital hypopituitarism (CH) is not entirely elucidated. FGFR1 and PROKR2 loss-of-function mutations are classically involved in hypogonadotrophic hypogonadism (HH), however, due to the clinical and genetic overlap of HH and CH; these genes may also be involved in the pathogenesis of CH. Using a candidate gene approach, we screened 156 Brazilian patients with combined pituitary hormone deficiencies (CPHD) for loss-of-function mutations in FGFR1 and PROKR2. We identified three FGFR1 variants (p.Arg448Trp, p.Ser107Leu and p.Pro772Ser) in four unrelated patients (two males) and two PROKR2 variants (p.Arg85Cys and p.Arg248Glu) in two unrelated female patients. Five of the six patients harbouring the variants had a first-degree relative that was an unaffected carrier of it. Results of functional studies indicated that the new FGFR1 variant p.Arg448Trp is a loss-of-function variant, while p.Ser107Leu and p.Pro772Ser present signalling activity similar to the wild-type form. Regarding PROKR2 variants, results from previous functional studies indicated that p.Arg85Cys moderately compromises receptor signalling through both MAPK and Ca(2) (+) pathways while p.Arg248Glu decreases calcium mobilization but has normal MAPK activity. The presence of loss-of-function variants of FGFR1 and PROKR2 in our patients with CPHD is indicative of an adjuvant and/or modifier effect of these rare variants on the phenotype. The presence of the same variants in unaffected relatives implies that they cannot solely cause the phenotype. Other associated genetic and/or environmental modifiers may play a role in the aetiology of this condition

Mutations in the SRY, DAX1, SF1 and WNT4 genes in Brazilian sex-reversed patients

In most mammals, male development is triggered by the transient expression of the SRY gene, which initiates a cascade of gene interactions ultimately leading to the formation of a testis from the indifferent fetal gonad. Mutation studies have identified several genes essential for early gonadal development. We report here a molecular study of the SRY, DAX1, SF1 and WNT4 genes, mainly involved in sexual determination, in Brazilian 46,XX and 46,XY sex-reversed patients. The group of 46,XX sex-reversed patients consisted of thirteen 46,XX true hermaphrodites and four 46,XX males, and was examined for the presence of the SRY gene and for the loss of function (inactivating mutations and deletions) of DAX1 and WNT4 genes. In the second group consisting of thirty-three 46,XY sex-reversed patients we investigated the presence of inactivating mutations in the SRY and SF1 genes as well as the overexpression (duplication) of the DAX1 and WNT4 genes. The SRY gene was present in two 46,XX male patients and in none of the true hermaphrodites. Only one mutation, located outside homeobox domain of the 5' region of the HMG box of SRY (S18N), was identified in a patient with 46,XY sex reversal. A novel 8-bp microdeletion of the SF1 gene was identified in a 46,XY sex-reversed patient without adrenal insufficiency. The dosage of DAX1 and WNT4 was normal in the sex-reversed patients studied. We conclude that these genes are rarely involved in the etiology of male gonadal development in sex-reversed patients, a fact suggesting the presence of other genes in the sex determination cascade

Analysis Of Anti-müllerian Hormone (amh) And Its Receptor (amhr2) Genes In Patients With Persistent Müllerian Duct Syndrome [pesquisa De Mutações Nos Genes Do Hormônio Antimülleriano (amh) E Do Seu Receptor (amhr2) Em Pacientes Com Síndrome De Persistência Dos Ductos Müllerianos]

Objective: To screen for mutations in AMH and AMHR2 genes in patients with persistent Müllerian duct syndrome (PMDS). Patients and method: Genomic DNA of eight patients with PMDS was obtained from peripheral blood leukocytes. Directed sequencing of the coding regions and the exon-intron boundaries of AMH and AMHR2 were performed. Results: The AMH mutations p.Arg95*, p.Arg123Trp, c.556-2A>G, and p.Arg502Leu were identified in five patients; and p.Gly323Ser and p.Arg407* in AMHR2 of two individuals. In silico analyses of the novel c.556-2A>G, p.Arg502Leu and p.Arg407* mutations predicted that they were harmful and were possible causes of the disease. Conclusion: A likely molecular etiology was found in the eight evaluated patients with PMDS. Four mutations in AMH and two in AMHR2 were identified. Three of them are novel mutations, c.556-2A>G, and p.Arg502Leu in AMH; and p.Gly323Ser in AMHR2. © ABE&M todos os direitos reservados.568473478Jost, A., Problems of fetal endocrinology: The gonadal and hypophyseal hormones (1953) Recent Prog Horm Res, 8, pp. 379-418Farikullah, J., Ehtisham, S., Nappo, S., Patel, L., Hennayake, S., Persistent Müllerian duct syndrome: Lessons learned from managing a series of eight patients over a 10-year period and review of literature regarding malignant risk from the Mullerian remnants (2012) BJU Int, , Epub 2012 Apr 30Rey, R., Picard, J.Y., Josso, N., Síndrome de Persistência dos Ductos de Müller (2010) Menino ou Menina? Distúrbios da Diferenciação do Sexo, pp. 279-295. , In: Guerra ATM, Guerra Júnior G, editors, 2. ed. RubioHutson, J.M., Davidson, P.M., Reece, L.A., Baker, M., Zhou, B., Failure of gubernacular development in the persistent Müllerian duct syndrome allows herniation of the testes (1994) Pediatr Surg Int, 9, pp. 544-546Abduljabbar, M., Taheini, K., Picard, J.Y., Cate, R.L., Josso, N., Mutations of the AMH Type II receptor in two extended families with persistent Müllerian duct syndrome: Lack of phenotype/genotype correlation (2012) Horm Res Paediatr, 77 (5), pp. 291-297Imbeaud, S., Faure, E., Lamarre, I., Mattei, M.G., Di Clemente, N., Tizard, R., Insensitivity to anti-Müllerian hormone due to a mutation in the human anti-Müllerian hormone receptor (1995) Nat Genet, 11 (4), pp. 382-388Cate, R.L., Mattaliano, R.J., Hession, C., Tizard, R., Farber, N.M., Cheung, A., Isolation of the bovine and human genes for Müllerian inhibiting substance and expression of the human gene in animal cells (1986) Cell, 45 (5), pp. 685-698Josso, N., Di Clemente, N., TGF-beta family members and gonadal development (1999) Trends Endocrinol Metab, 10 (6), pp. 216-222Josso, N., Belville, C., Di Clemente, N., Picard, J.Y., AMH and AMH receptor defects in persistent Müllerian duct syndrome (2005) Hum Reprod Update, 11 (4), pp. 351-356Menabo, S., Balsamo, A., Nicoletti, A., Gennari, M., Pirazzoli, P., Cicognani, A., Three novel AMH gene mutations in a patient with persistent Müllerian duct syndrome and normal AMH serum dosage (2008) Horm Res, 70 (2), pp. 124-128van der Zwan, Y.G., Bruggenwirth, H.T., Drop, S.L., Wolffenbuttel, K.P., Madern, G.C., Looijenga, L.H., A novel AMH missense mutation in a patient with persistent Müllerian duct syndrome (2012) Sex Dev, , Epub 2012 Jul 11Baarends, W.M., van Helmond, M.J., Post, M., van der Schoot, P.J., Hoogerbrugge, J.W., de Winter, J.P., A novel member of the transmembrane serine/threonine kinase receptor family is specifically expressed in the gonads and in mesenchymal cells adjacent to the Müllerian duct (1994) Development, 120 (1), pp. 189-197Di Clemente, N., Wilson, C., Faure, E., Boussin, L., Carmillo, P., Tizard, R., Cloning, expression, and alternative splicing of the receptor for anti-Müllerian hormone (1994) Mol Endocrinol, 8 (8), pp. 1006-1020Belville, C., Marechal, J.D., Pennetier, S., Carmillo, P., Masgrau, L., Messika-Zeitoun, L., Natural mutations of the anti-Müllerian hormone type II receptor found in persistent Müllerian duct syndrome affect ligand binding, signal transduction and cellular transport (2009) Hum Mol Genet, 18 (16), pp. 3002-3013Miller, S.A., Dykes, D.D., Polesky, H.F., A simple salting out procedure for extracting DNA from human nucleated cells (1988) Nucleic Acids Res, 16 (3), p. 1215Imbeaud, S., Carre-Eusebe, D., Rey, R., Belville, C., Josso, N., Picard, J.Y., Molecular genetics of the persistent Müllerian duct syndrome: A study of 19 families (1994) Hum Mol Genet, 3 (1), pp. 125-131Josso, N., Picard, J.Y., Rey, R., Di Clemente, N., Testicular anti-Müllerian hormone: History, genetics, regulation and clinical applications (2006) Pediatr Endocrinol Rev, 3 (4), pp. 347-358Rey, R.A., Codner, E., Iniguez, G., Bedecarras, P., Trigo, R., Okuma, C., Low risk of impaired testicular Sertoli and Leydig cell functions in boys with isolated hypospadias (2005) J Clin Endocrinol Metab, 90 (11), pp. 6035-6040Imbeaud, S., Belville, C., Messika-Zeitoun, L., Rey, R., Di Clemente, N., Josso, N., A 27 base-pair deletion of the anti-Müllerian type II receptor gene is the most common cause of the persistent Müllerian duct syndrome (1996) Hum Mol Genet, 5 (9), pp. 1269-127

The Sitting Height/height Ratio For Age In Healthy And Short Individuals And Its Potential Role In Selecting Short Children For Shox Analysis

Aims: To determine the presence of abnormal body proportion, assessed by sitting height/height ratio for age and sex (SH/H SDS) in healthy and short individuals, and to estimate its role in selecting short children for SHOX analysis. Methods: Height, sitting height and weight were evaluated in 1,771 healthy children, 128 children with idiopathic short stature (ISS), 58 individuals with SHOX defects (SHOX-D) and 193 females with Turner syndrome (TS). Results: The frequency of abnormal body proportion, defined as SH/H SDS >2, in ISS children was 16.4% (95% CI 10-22%), which was higher than in controls (1.4%, 95% CI 0.8-1.9%, p 2 were less common in children (48%, 95% CI 37-59%) and in adults (28%, 95% CI 20-36%) with TS. Conclusion: Abnormal body proportions were observed in almost all individuals with SHOX-D, 50% of females with TS and 16% of children considered ISS. Defects in SHOX gene were identified in 19% of ISS children with SH/H SDS >2, suggesting that SH/H SDS is a useful tool to select children for undergoing SHOX molecular studies. © 2013 S. Karger AG, Basel.806449456Ellison, J.W., Wardak, Z., Young, M.F., Gehron Robey, P., Laig-Webster, M., Chiong, W., PHOG, a candidate gene for involvement in the short stature of turner syndrome (1997) Hum Mol Genet, 6, pp. 1341-1347Rao, E., Weiss, B., Fukami, M., Rump, A., Niesler, B., Mertz, A., Muroya, K., Rappold, G.A., Pseudoautosomal deletions encompassing a novel homeobox gene cause growth failure in idiopathic short stature and Turner syndrome (1997) Nat Genet, 16, pp. 54-63Clement-Jones, M., Schiller, S., Rao, E., Blaschke, R.J., Zuniga, A., Zeller, R., Robson, S.C., Rappold, G.A., The short stature homeobox gene SHOX is involved in skeletal abnormalities in Turner syndrome (2000) Hum Mol Genet, 9, pp. 695-702Ross, J.L., Kowal, K., Quigley, C.A., Blum, W.F., Cutler Jr., G.B., Crowe, B., Hovanes, K., Zinn, A.R., The phenotype of short stature homeobox gene (SHOX) deficiency in childhood: Contrasting children with Leri-Weill dyschondrosteosis and Turner syndrome (2005) J Pediatr, 147, pp. 499-507Neufeld, N.D., Lippe, B.M., Kaplan, S.A., Disproportionate growth of the lower extremities. A major determinant of short stature in Turner's syndrome (1978) Am J Dis Child, 132, pp. 296-298Rongen-Westerlaken, C., Rikken, B., Vastrick, P., Jeuken, A.H., De Lange, M.Y., Wit, J.M., Van Der Tweel, L., Van Den Brande, J.L., Body proportions in individuals with Turner syndrome The Dutch growth hormone working group (1993) Eur J Pediatr, 152, pp. 813-817Gravholt, C.H., Weis Naeraa, R., Reference values for body proportions and body composition in adult women with Ullrich-Turner syndrome (1997) Am J Med Genet, 72, pp. 403-408Baldin, A.D., Armani, M.C., Morcillo, A.M., Lemos-Marini, S.H., Baptista, M.T., Maciel-Guerra, A.T., Guerra-Junior, G., Body proportions in a group of Brazilian patients with Turner syndrome (2005) Arq Bras Endocrinol Metabol, 49, pp. 529-535Jorge, A.A., Funari, M.F., Nishi, M.Y., Mendonca, B.B., Short stature caused by isolated SHOX gene haploinsufficiency: Update on the diagnosis and treatment (2010) Pediatr Endocrinol Rev, 8, pp. 79-85Ross, J.L., Scott Jr., C., Marttila, P., Kowal, K., Nass, A., Papenhausen, P., Abboudi, J., Zinn, A.R., Phenotypes associated with SHOX deficiency (2001) J Clin Endocrinol Metab, 86, pp. 5674-5680Binder, G., Ranke, M.B., Martin, D.D., Auxology is a valuable instrument for the clinical diagnosis of SHOX haploinsufficiency in schoolage children with unexplained short stature (2003) J Clin Endocrinol Metab, 88, pp. 4891-4896Rappold, G., Blum, W.F., Shavrikova, E.P., Crowe, B.J., Roeth, R., Quigley, C.A., Ross, J.L., Niesler, B., Genotypes and phenotypes in children with short stature: Clinical indicators of SHOX haploinsufficiency (2007) J Med Genet, 44, pp. 306-313Jorge, A.A., Souza, S.C., Nishi, M.Y., Billerbeck, A.E., Liborio, D.C., Kim, C.A., Arnhold, I.J., Mendonca, B.B., SHOX mutations in idiopathic short stature and Leri-Weill dyschondrosteosis: Frequency and phenotypic variability (2007) Clin Endocrinol (Oxf), 66, pp. 130-135Hirschfeldova, K., Solc, R., Baxova, A., Zapletalova, J., Kebrdlova, V., Gaillyova, R., Prasilova, S., Stekrova, J., SHOX gene defects and selected dysmorphic signs in patients of idiopathic short stature and Leri-Weill dyschondrosteosis (2012) Gene, 491, pp. 123-127Kant, S.G., Broekman, S.J., De Wit, C.C., Bos, M., Scheltinga, S.A., Bakker, E., Oostdijk, W., Losekoot, M., Phenotypic characterization of patients with deletions in the 3 ′ flanking SHOX region (2013) PeerJ, 1, pp. e35Fredriks, A.M., Van Buuren, S., Van Heel, W.J., Dijkman-Neerincx, R.H., Verloove-Vanhorick, S.P., Wit, J.M., Nationwide age references for sitting height, leg length, and sitting height/ height ratio, and their diagnostic value for disproportionate growth disorders (2005) Arch Dis Child, 90, pp. 807-812Kuczmarski, R.J., Ogden, C.L., Grummer-Strawn, L.M., Flegal, K.M., Guo, S.S., Wei, R., Mei, Z., Johnson, C.L., CDC growth charts: United States (2000) Adv Data, 314, pp. 1-27Silva, D.A., Pelegrini, A., Petroski, E.L., Gaya, A.C., Comparison between the growth of Brazilian children and adolescents and the reference growth charts: Data from a Brazilian project (2010) J Pediatr (Rio J), 86, pp. 115-120Funari, M.F., Jorge, A.A., Souza, S.C., Billerbeck, A.E., Arnhold, I.J., Mendonca, B.B., Nishi, M.Y., Usefulness of MLPA in the detection of SHOX deletions (2010) Eur J Med Genet, 53, pp. 234-238Lango Allen, H., Estrada, K., Lettre, G., Berndt, S.I., Weedon, M.N., Rivadeneira, F., Willer, C.J., Hirschhorn, J.N., Hundreds of variants clustered in genomic loci and biological pathways affect human height (2010) Nature, 467, pp. 832-838Shears, D.J., Vassal, H.J., Goodman, F.R., Palmer, R.W., Reardon, W., Superti-Furga, A., Scambler, P.J., Winter, R.M., Mutation and deletion of the pseudoautosomal gene SHOX cause Leri-Weill dyschondrosteosis (1998) Nat Genet, 19, pp. 70-73Belin, V., Cusin, V., Viot, G., Girlich, D., Toutain, A., Moncla, A., Vekemans, M., Cormier-Daire, V., SHOX mutations in dyschondrosteosis (Leri-Weill syndrome) (1998) Nat Genet, 19, pp. 67-69Rappold, G.A., Fukami, M., Niesler, B., Schiller, S., Zumkeller, W., Bettendorf, M., Heinrich, U., Ogata, T., Deletions of the homeobox gene SHOX (short stature homeobox) are an important cause of growth failure in children with short stature (2002) J Clin Endocrinol Metab, 87, pp. 1402-1406Jorge, A.A., Arnhold, I.J., Anthropometric evaluation of children with SHOX mutations can be used as indication for genetic studies in children of short stature (2007) J Med Genet, 44, pp. e90. , author reply e91Kosho, T., Muroya, K., Nagai, T., Fujimoto, M., Yokoya, S., Sakamoto, H., Hirano, T., Ogata, T., Skeletal features and growth patterns in 14 patients with haploinsufficiency of SHOX: Implications for the development of turner syndrome (1999) J Clin Endocrinol Metab, 84, pp. 4613-4621Fukami, M., Nishi, Y., Hasegawa, Y., Miyoshi, Y., Okabe, T., Haga, N., Nagai, T., Ogata, T., Statural growth in 31 Japanese patients with SHOX haploinsufficiency: Support for a disadvantageous effect of gonadal estrogens (2004) Endocr J, 51, pp. 197-200Blum, W.F., Cao, D., Hesse, V., Fricke-Otto, S., Ross, J.L., Jones, C., Quigley, C.A., Binder, G., Height gains in response to growth hormone treatment to final height are similar in patients with SHOX deficiency and Turner syndrome (2009) Horm Res, 71, pp. 167-172Scalco, R.C., Melo, S.S., Pugliese-Pires, P.N., Funari, M.F., Nishi, M.Y., Arnhold, I.J., Mendonca, B.B., Jorge, A.A., Effectiveness of the combined recombinant human growth hormone and gonadotropin-releasing hormone analog therapy in pubertal patients with short stature due to SHOX deficiency (2010) J Clin Endocrinol Metab, 95, pp. 328-33

Supplementary Material for: GH-Releasing Hormone Receptor Gene: A Novel Splice-Disrupting Mutation and Study of Founder Effects

<b><i>Background:</i></b> Mutations in GH-releasing hormone receptor gene <i>(GHRHR)</i> are emerging as the most common cause of autosomal recessive isolated GH deficiency (IGHD). <b><i>Objective:</i></b> To search for <i>GHRHR</i> mutations in patients with familial or sporadic IGHD and to investigate founder effects in recurring mutations. <b><i>Methods:</i></b> The coding region of <i>GHRHR</i> was entirely amplified and sequenced from DNA of 18 patients with IGHD (16 unrelated) with topic posterior pituitary lobe on MRI. Haplotypes containing promoter SNPs and microsatellites flanking <i>GHRHR</i> were analyzed in patients with c.57+1G>A (IVS1+1G>A) mutation of our previously published kindred and also a Brazilian patient and 2 previously reported Japanese sisters with c.1146G>A (p.E382E) mutation. <b><i>Results:</i></b> A novel homozygous intronic <i>GHRHR</i> c.752-1G>A (IVS7-1G>A) mutation, predicting loss of the constitutive splice acceptor site, was identified in two siblings with IGHD. A compound heterozygous c.[57+1G>A];[1146G>A] and a heterozygous c.527C>T (p.A176V) were found in two sporadic cases. Haplotype analysis provided evidence for a founder effect for the c.57+1G>A mutation and independent recurrence for the c.1146G>A mutation. <b><i>Conclusion:</i></b> We report a novel splice-disrupting mutation in <i>GHRHR</i> in 2 siblings and provide evidence that all c.57+1G>A (IVS1+1G>A) mutant chromosomes have the same haplotype ancestor, indicating the occurrence of a founder effect in Brazilian patients with IGHD

Mitochondrial functions on oocytes and preimplantation embryos*

Oocyte quality has long been considered as a main limiting factor for in vitro fertilization (IVF). In the past decade, extensive observations demonstrated that the mitochondrion plays a vital role in the oocyte cytoplasm, for it can provide adenosine triphosphate (ATP) for fertilization and preimplantation embryo development and also act as stores of intracellular calcium and proapoptotic factors. During the oocyte maturation, mitochondria are characterized by distinct changes of their distribution pattern from being homogeneous to heterogeneous, which is correlated with the cumulus apoptosis. Oocyte quality decreases with the increasing maternal age. Recent studies have shown that low quality oocytes have some age-related dysfunctions, which include the decrease in mitochondrial membrane potential, increase of mitochondrial DNA (mtDNA) damages, chromosomal aneuploidies, the incidence of apoptosis, and changes in mitochondrial gene expression. All these dysfunctions may cause a high level of developmental retardation and arrest of preimplantation embryos. It has been suggested that these mitochondrial changes may arise from excessive reactive oxygen species (ROS) that is closely associated with the oxidative energy production or calcium overload, which may trigger permeability transition pore opening and subsequent apoptosis. Therefore, mitochondria can be seen as signs for oocyte quality evaluation, and it is possible that the oocyte quality can be improved by enhancing the physical function of mitochondria. Here we reviewed recent advances in mitochondrial functions on oocytes