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1.
Minerva Endocrinol ; 38(1): 1-16, 2013 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-23435439

RESUMO

Growth hormone (GH) deficiency (GHD) in humans manifests differently according to the individual developmental stage (early after birth, during childhood, at puberty or in adulthood), the cause or mechanism (genetic, acquired or idiopathic), deficiency intensity and whether it is the only pituitary-affected hormone or is combined with that of other pituitary hormones or forms part of a complex syndrome. Growing knowledge of the genetic basis of GH deficiency continues to provide us with useful information to further characterise mutation types and mechanisms for previously described and new candidate genes. Despite these advances, a high proportion of GH deficiencies with no recognisable acquired basis continue to be labelled as idiopathic, although less frequently when they are congenital and/or familial. The clinical and biochemical diagnoses continue to be a conundrum despite efforts to harmonise biochemical assays for GH and IGF-1 analysis, probably because the diagnosis based on the so-called GH secretion stimulation tests will prove to be of limited usefulness for predicting therapy indications.


Assuntos
Hormônio do Crescimento Humano/deficiência , Hormônio do Crescimento Humano/genética , Mutação , Adolescente , Desenvolvimento do Adolescente , Biomarcadores/sangue , Estatura/efeitos dos fármacos , Criança , Desenvolvimento Infantil , Nanismo Hipofisário/genética , Hormônio do Crescimento Humano/sangue , Humanos , Fator de Crescimento Insulin-Like I/metabolismo , Fenótipo , Puberdade/efeitos dos fármacos
2.
Int J Androl ; 34(6 Pt 2): e526-35, 2011 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-21631525

RESUMO

One hundred and forty-six index patients with 46,XY DSD in whom gonads were confirmed as testes were consecutively studied for a molecular diagnosis during the period 2002-2010. AR gene was analysed in all patients as the first candidate gene, yielding a mutation in 42.5% of cases and SRD5A2 gene was analysed as the second candidate gene, resulting in the characterization of 10 different mutations (p.Y91D, p.G115D, p.Q126R, p.R171S, p.Y188CfsX9, p.N193S, p.A207D, p.F219SfsX60, p.R227Q and p.R246W) in nine index patients (6.2% of the total number of 46,XY DSD patients). One of the mutations (p.Y188CfsX9) has never been reported. In addition, we genotyped SRD5A2 gene p.V89L and c.281+15T>C polymorphisms in 46,XY DSD and in 156 normal adult males and found that patients with SRD5A2 mutations or without a known molecular diagnosis presented a higher frequency of homozygous p.L89, homozygous TT and combined CCTT genotypes compared with controls. This result suggests that 46,XY DSD patient phenotypes may be influenced by SRD5A2 polymorphism genotypes. SRD5A2 gene mutations may not be as infrequent as previously considered in 46,XY DSD patients with variable degrees of external genitalia virilization at birth and normal T production and appears to be the second aetiology in our series.


Assuntos
3-Oxo-5-alfa-Esteroide 4-Desidrogenase/genética , Transtornos do Desenvolvimento Sexual/genética , Proteínas de Membrana/genética , Mutação , Polimorfismo de Nucleotídeo Único , Sequência de Bases , Primers do DNA , Humanos , Reação em Cadeia da Polimerase , Espanha
3.
J Clin Endocrinol Metab ; 93(3): 764-70, 2008 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-18160465

RESUMO

CONTEXT: Consensus is lacking as to whether the exon 3-deleted (d3)/full-length (fl) GH receptor (GHR) polymorphism is associated with responsiveness to GH therapy. OBJECTIVE: Our objective was to evaluate, in short, prepubertal, appropriate-for-gestational age (AGA) patients, 2-yr growth response to GH therapy (31.7+/-3.5 microg/kg.d) according to exon 3-deleted/full-length GHR genotypes. DESIGN: We conducted a retrospective study. PATIENTS: We studied 106 short AGA children, 58 boys and 48 girls, 7.8+/-2.3 yr, (d3/d3 n=18, d3/fl n=42, and fl/fl n=46). The GH response to two provocative stimuli were under 10 ng/ml in 65 and one or both over 10 ng/ml in 41 patients. MAIN OUTCOME MEASURES: Patients were followed by a single clinical team and remained prepubertal during the study. The exon 3-deleted/full-length GHR genotypes were determined and analyzed in the same hospital. RESULTS: Growth velocity significantly (P<0.0001) increased during the first and second years of therapy, as did height sd score (SDS). These increases were similar in each exon 3-deleted/full-length GHR genotype. Total 2-yr height gain (SDS) did not differ statistically among genotypes: 15.5+/-2.2 cm and 1.2+/-0.5 SDS in d3/d3, 15.9+/-2.0 cm and 1.3+/-0.4 SDS in d3/fl, and 15.4+/-2.1 cm and 1.1+/-0.3 SDS in fl/fl. No significant differences among the three genotypes were found in both sexes or in patients with different GH peak response to provocative stimuli for these parameters. An analysis of previously published studies was also performed. CONCLUSIONS: These results confirm in AGA patients those previously found by us and others in small-for-gestational-age patients and suggest that neither sex nor GH peaks after provocative stimuli might influence significantly the responsiveness to GH therapy according to the exon 3-deleted/full-length GHR genotypes.


Assuntos
Estatura/efeitos dos fármacos , Éxons , Transtornos do Crescimento/genética , Hormônio do Crescimento/uso terapêutico , Terapia de Reposição Hormonal , Polimorfismo Genético , Receptores da Somatotropina/genética , Peso ao Nascer , Criança , Pré-Escolar , Feminino , Genótipo , Transtornos do Crescimento/tratamento farmacológico , Hormônio do Crescimento Humano/sangue , Humanos , Recém-Nascido , Masculino , Estudos Retrospectivos
4.
J Clin Endocrinol Metab ; 93(7): 2709-15, 2008 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-18445665

RESUMO

CONTEXT: The exon 3-deleted/full-length (d3/fl) GH receptor polymorphism (d3/fl-GHR) has been associated with responsiveness to GH therapy in short small-for-gestational-age (SGA) patients, although consensus is lacking. However, its influence on glucose homeostasis, at baseline or under GH therapy, has not been investigated. OBJECTIVE: Our objective was to evaluate whether the d3/fl-GHR genotypes influence insulin sensitivity in short SGA children before or after puberty onset or during GH therapy. DESIGN: We conducted a 2-yr prospective, controlled, randomized trial. SETTING: Thirty Spanish hospitals participated. Auxological, GH secretion, and glucose homeostasis evaluation was hospital based, whereas molecular analyses and data computation were centralized. PATIENTS: Patients included 219 short SGA children [body mass index sd score (SDS) < or = 2.0]; 159 were prepubertal (group 1), and 60 had entered puberty (group 2). INTERVENTION: Seventy-eight patients from group 1 were treated with GH (66 microg/kg.d) for 2 yr (group 3). MAIN OUTCOME MEASURES: Previous and 2-yr follow-up auxological and biochemical data were recorded, d3/fl-GHR genotypes determined, and data analyzed. RESULTS: In groups 1 and 2, fasting glucose, insulin, homeostasis model assessment (HOMA), and quantitative insulin sensitivity check index (QUICKI) were similar in each d3/fl-GHR genotype. Group 2 glucose, insulin, and HOMA were significantly higher and QUICKI lower than in group 1. In group 3 GH-treated patients, height SDS, growth velocity SDS, fasting glucose, insulin, and HOMA significantly increased as did body mass index SDS at the end of the second year, and QUICKI decreased during the first and second years, with no differences among the d3/fl-GHR genotypes. CONCLUSION: In short SGA patients, the d3/fl-GHR genotypes do not seem to influence prepubertal or pubertal insulin sensitivity indexes or their changes over 2 yr of GH therapy (66 mug/kg.d).


Assuntos
Glucose/metabolismo , Hormônio do Crescimento Humano/uso terapêutico , Recém-Nascido Pequeno para a Idade Gestacional , Polimorfismo Genético , Puberdade , Receptores da Somatotropina/genética , Índice de Massa Corporal , Criança , Éxons , Feminino , Deleção de Genes , Homeostase , Hormônio do Crescimento Humano/deficiência , Humanos , Recém-Nascido , Masculino , Estudos Prospectivos
5.
Eur J Endocrinol ; 179(4): R197-R206, 2018 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-30299888

RESUMO

The differential diagnosis of differences or disorders of sex development (DSD) belongs to the most complex fields in medicine. It requires a multidisciplinary team conducting a synoptic and complementary approach consisting of thorough clinical, hormonal and genetic workups. This position paper of EU COST (European Cooperation in Science and Technology) Action BM1303 'DSDnet' was written by leading experts in the field and focuses on current best practice in genetic diagnosis in DSD patients. Ascertainment of the karyotpye defines one of the three major diagnostic DSD subclasses and is therefore the mandatory initial step. Subsequently, further analyses comprise molecular studies of monogenic DSD causes or analysis of copy number variations (CNV) or both. Panels of candidate genes provide rapid and reliable results. Whole exome and genome sequencing (WES and WGS) represent valuable methodological developments that are currently in the transition from basic science to clinical routine service in the field of DSD. However, in addition to covering known DSD candidate genes, WES and WGS help to identify novel genetic causes for DSD. Diagnostic interpretation must be performed with utmost caution and needs careful scientific validation in each DSD case.


Assuntos
Transtornos do Desenvolvimento Sexual/diagnóstico , Sequenciamento do Exoma , Cariótipo , Sequenciamento Completo do Genoma , Hiperplasia Suprarrenal Congênita/diagnóstico , Hiperplasia Suprarrenal Congênita/genética , Variações do Número de Cópias de DNA , Transtornos do Desenvolvimento Sexual/genética , União Europeia , Disgenesia Gonadal/diagnóstico , Disgenesia Gonadal/genética , Humanos , Biologia Molecular , Técnicas de Diagnóstico Molecular , Guias de Prática Clínica como Assunto , Análise de Sequência de DNA
6.
J Clin Endocrinol Metab ; 91(9): 3281-6, 2006 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-16804042

RESUMO

CONTEXT: The d3/fl-GH receptor (d3/fl-GHR, exon 3-deleted/full-length GHR) has recently been associated with responsiveness to GH therapy. OBJECTIVE: The objective of the study was to evaluate whether the d3/fl-GHR genotypes influence the intensity of spontaneous and/or GH therapy-stimulated growth in small-for-gestational-age (SGA) patients. DESIGN: This was a 2-yr prospective, controlled, randomized trial. SETTING: Thirty Spanish hospitals participated. Auxologic and GH secretion evaluation was hospital based, whereas molecular analyses and auxologic data computation were centralized. PATIENTS: Patients included 170 short SGA children: 140 remained prepubertal and 30 entered puberty during the second follow-up year. INTERVENTION: Eighty-six were treated with GH (66 microg/kg.d) for 2 yr and 84 were not treated. MAIN OUTCOME MEASURES: Previous and 2-yr follow-up auxologic data were recorded at each hospital, d3/fl-GHR genotypes determined, and data analyzed for patients who remained prepubertal (group 1, 68 GH treated and 72 non-GH treated) and for all the patients (group 2). RESULTS: In group 1 GH-treated patients, growth velocity, and height-sd score during the first and second years, total 2-yr height gain (18.5 +/- 2.4 cm in d3/d3; 18.4 +/- 2.6 in d3/fl; 19.5 +/- 2.3 in fl/fl), Delta 2-yr height increase (9.1 +/- 2.4 cm in d3/d3; 9.4 +/- 3.0 in d3/fl; 10.4 +/- 2.1 in fl/fl), first-year growth prediction and studentized residual values (0.08 +/- 1.26 in d3/d3; 0.28 +/- 1.21 in d3/fl; 0.67 +/- 0.95 in fl/fl) did not differ among the d3/fl-GHR genotypes. In group 1 non-GH-treated patients, neither growth velocity nor height-sd score changed significantly, and values were similar in each d3/fl-GHR genotype. Results in all patients (group 2) were similar to those in group 1. CONCLUSIONS: In short non-GH-deficient SGA children, both spontaneous growth rate and responsiveness to 66 microg/k.d GH therapy were similar for each d3/fl-GHR genotype carried.


Assuntos
Estatura/efeitos dos fármacos , Hormônio do Crescimento Humano/uso terapêutico , Receptores da Somatotropina/genética , Criança , Método Duplo-Cego , Feminino , Genótipo , Hormônio do Crescimento Humano/sangue , Hormônio do Crescimento Humano/metabolismo , Humanos , Recém-Nascido , Proteína 3 de Ligação a Fator de Crescimento Semelhante à Insulina/sangue , Fator de Crescimento Insulin-Like I/metabolismo , Masculino , Polimorfismo de Nucleotídeo Único , Estudos Prospectivos , Espanha , Estatísticas não Paramétricas
7.
J Clin Endocrinol Metab ; 101(11): 4468-4477, 2016 11.
Artigo em Inglês | MEDLINE | ID: mdl-27583472

RESUMO

CONTEXT: Only approximately 85% of patients with a clinical diagnosis complete androgen insensitivity syndrome and less than 30% with partial androgen insensitivity syndrome can be explained by inactivating mutations in the androgen receptor (AR) gene. OBJECTIVE: The objective of the study was to clarify this discrepancy by in vitro determination of AR transcriptional activity in individuals with disorders of sex development (DSD) and male controls. DESIGN: Quantification of DHT-dependent transcriptional induction of the AR target gene apolipoprotein D (APOD) in cultured genital fibroblasts (GFs) (APOD assay) and next-generation sequencing of the complete coding and noncoding AR locus. SETTING: The study was conducted at a university hospital endocrine research laboratory. PATIENTS: GFs from 169 individuals were studied encompassing control males (n = 68), molecular defined DSD other than androgen insensitivity syndrome (AIS; n = 18), AR mutation-positive AIS (n = 37), and previously undiagnosed DSD including patients with a clinical suspicion of AIS (n = 46). INTERVENTION(S): There were no interventions. MAIN OUTCOME MEASURE(S): DHT-dependent APOD expression in cultured GF and AR mutation status in 169 individuals was measured. RESULTS: The APOD assay clearly separated control individuals (healthy males and molecular defined DSD patients other than AIS) from genetically proven AIS (cutoff < 2.3-fold APOD-induction; 100% sensitivity, 93.3% specificity, P < .0001). Of 46 DSD individuals with no AR mutation, 17 (37%) fell below the cutoff, indicating disrupted androgen signaling. CONCLUSIONS: AR mutation-positive AIS can be reliably identified by the APOD assay. Its combination with next-generation sequencing of the AR locus uncovered an AR mutation-negative, new class of androgen resistance, which we propose to name AIS type II. Our data support the existence of cellular components outside the AR affecting androgen signaling during sexual differentiation with high clinical relevance.


Assuntos
Síndrome de Resistência a Andrógenos/diagnóstico , Apolipoproteínas D , Bioensaio/normas , Transtornos do Desenvolvimento Sexual/diagnóstico , Receptores Androgênicos/metabolismo , Testosterona/análogos & derivados , Adulto , Síndrome de Resistência a Andrógenos/genética , Síndrome de Resistência a Andrógenos/metabolismo , Células Cultivadas , Transtornos do Desenvolvimento Sexual/genética , Transtornos do Desenvolvimento Sexual/metabolismo , Fibroblastos , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Masculino , Mutação , Receptores Androgênicos/genética , Sensibilidade e Especificidade , Testosterona/metabolismo , Transcrição Gênica
8.
J Bone Miner Res ; 15(3): 534-40, 2000 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-10750568

RESUMO

Angiogenesis is a crucial event in endochondral ossification. Chemoattractants and mitogens for endothelial cells (such as basic fibroblast growth factor [bFGF] and transforming growth factor beta [TGF-beta]), which act as local regulators of the process, are synthesized by chondrocytes under several stimuli and in relation to the differentiation stage of the cartilage. Vascular endothelial growth factor (VEGF) is a 44-kDa protein well known as a potent angiogenic molecule owing to its mitogenic and permeability-causing properties. In this work, VEGF was located by immunohistochemistry in growth plate cartilage of human fetuses (20-22 weeks old) and its expression was demonstrated by reverse-transcription polymerase chain reaction (RT-PCR). Primary culture of human fetal epiphyseal chondrocytes (HFEC) maintained VEGF expression at protein and messenger RNA (mRNA) levels and this expression was stimulated by cartilage-promoting growth factors incorporated into the culture media (rFGF-b, rTGF-beta1, and insulin-like growth factor [rFGF-b] at 50 ng/ml). The conditioned medium (CM) of HFEC stimulated the proliferation of endothelial cells, and this was partially blocked by anti-VEGF antibody. These studies showed VEGF production by chondrocytes of the epiphyseal growth cartilage and suggested a role of this factor in cartilage physiology and the angiogenic process.


Assuntos
Cartilagem/embriologia , Fatores de Crescimento Endotelial/biossíntese , Proteínas Fetais/biossíntese , Lâmina de Crescimento/embriologia , Linfocinas/biossíntese , Neovascularização Fisiológica/fisiologia , Processamento Alternativo , Calcitriol/farmacologia , Cartilagem/citologia , Cartilagem/metabolismo , Linhagem Celular Transformada/efeitos dos fármacos , Células Cultivadas/metabolismo , Meios de Cultivo Condicionados , Replicação do DNA/efeitos dos fármacos , Fatores de Crescimento Endotelial/genética , Endotélio Vascular/citologia , Fator de Crescimento Epidérmico/farmacologia , Proteínas Fetais/genética , Fator 2 de Crescimento de Fibroblastos/farmacologia , Regulação da Expressão Gênica no Desenvolvimento , Idade Gestacional , Lâmina de Crescimento/irrigação sanguínea , Lâmina de Crescimento/citologia , Lâmina de Crescimento/metabolismo , Humanos , Fator de Crescimento Insulin-Like I/farmacologia , Fator de Crescimento Insulin-Like II/farmacologia , Linfocinas/genética , RNA Mensageiro/biossíntese , Proteínas Recombinantes/farmacologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fator de Crescimento Transformador beta/farmacologia , Tretinoína/farmacologia , Fator A de Crescimento do Endotélio Vascular , Fatores de Crescimento do Endotélio Vascular
9.
J Bone Miner Res ; 15(10): 2005-10, 2000 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-11028454

RESUMO

Lumbar L2-L4 bone mineral density (BMD) values were measured in 37 adolescent and young adult Turner syndrome patients. Nine had developed spontaneous puberty and had had regular menses since menarche (12.55 years +/- 1.17 years) to the time of BMD evaluation (14.96 years +/- 1.26 years). In the other 28, puberty was induced with increasing doses of oral ethinyl estradiol (2.5-10.0 microg/day, for 2 years) and later administration of estrogen/gestagen therapy up to the time of BMD evaluation. In 18, the adolescent group, menarche appeared at 14.68 years +/- 0.63 years and BMD was evaluated at 17.77 years +/- 0.70 years, and in the other 10, the young adult group, menarche appeared at 14.47 years +/- 0.53 years and BMD was evaluated at 20.90 years +/- 0.68 year. BMD values were in the normal range in those who had developed spontaneous puberty (Z score values, -0.24 +/- 0.22) and in the osteopenia range in those in whom puberty was induced (Z score values, -2.09 +/- 0.79 and -2.18 +/- 0.32 for the adolescent and young adult groups, respectively) p < 0.0001. Height Z score values were similar in all three groups (-3.45 +/- 0.77, -3.15 +/- 0.83, and -3.08 +/- 0.33, respectively). No significant differences in calcium intake or physical activity were found among groups. Neither the karyotype distribution nor growth hormone (GH) therapy (five in the spontaneous puberty and six in the induced puberty groups had been treated for a 3.5- to 4.4-year period) explained the differences in BMD values. Because the main difference between groups was the availability of estrogens to bone tissue from infancy to menarche and of estrogens/gestagens from then on up to the time of BMD evaluation, our results suggest that normal gonadal function from infancy to adulthood may be required for adequate bone mass peaking. Early detection of osteopenia and improvement in general measures for adequate bone mass peaking (calcium intake and physical activity) should be considered mandatory in the health care of these patients.


Assuntos
Densidade Óssea , Puberdade , Síndrome de Turner/fisiopatologia , Absorciometria de Fóton , Adolescente , Adulto , Envelhecimento/fisiologia , Estatura , Densidade Óssea/efeitos dos fármacos , Densidade Óssea/genética , Doenças Ósseas Metabólicas/complicações , Doenças Ósseas Metabólicas/diagnóstico , Cálcio/administração & dosagem , Etinilestradiol/farmacologia , Etinilestradiol/uso terapêutico , Exercício Físico , Feminino , Hormônio do Crescimento/uso terapêutico , Humanos , Cariotipagem , Vértebras Lombares/diagnóstico por imagem , Menarca/efeitos dos fármacos , Menarca/genética , Puberdade/efeitos dos fármacos , Puberdade/genética , Estatística como Assunto , Síndrome de Turner/complicações , Síndrome de Turner/tratamento farmacológico , Síndrome de Turner/genética
10.
J Bone Miner Res ; 15(11): 2063-73, 2000 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-11092389

RESUMO

We evaluated a 7-year-old girl with severe platelike osteoma cutis (POC), a variant of progressive osseous heteroplasia (POH). The child had congenital heterotopic ossification of dermis and subcutaneous fat that progressed to involve deep skeletal muscles of the face, scalp, and eyes. Although involvement of skeletal muscle is a prominent feature of POH, heterotopic ossification has not been observed in the head, face, or extraocular muscles. The cutaneous ossification in this patient was suggestive of Albright hereditary osteodystrophy (AHO); however, none of the other characteristic features of AHO were expressed. Inactivating mutations of the GNAS1 gene, which encodes the alpha-subunit of the stimulatory G protein of adenylyl cyclase, is the cause of AHO. Mutational analysis of GNAS1 using genomic DNA of peripheral blood and of lesional and nonlesional tissue from our patient revealed a heterozygous 4-base pair (bp) deletion in exon 7, identical to mutations that have been found in some AHO patients. This 4-bp deletion in GNAS1 predicts a protein reading frameshift leading to 13 incorrect amino acids followed by a premature stop codon. To investigate pathways of osteogenesis by which GNAS1 may mediate its effects, we examined the expression of the obligate osteogenic transcription factor Cbfa1/RUNX2 in lesional and uninvolved dermal fibroblasts from our patient and discovered expression of bone-specific Cbfa1 messenger RNA (mRNA) in both cell types. These findings document severe heterotopic ossification in the absence of AHO features caused by an inactivating GNAS1 mutation and establish the GNAS1 gene as the leading candidate gene for POH.


Assuntos
Subunidades alfa Gs de Proteínas de Ligação ao GTP/genética , Mutação , Proteínas de Neoplasias , Ossificação Heterotópica/genética , Ossificação Heterotópica/patologia , Fatores de Transcrição/genética , Sequência de Aminoácidos , Sequência de Bases , Osso e Ossos/metabolismo , Linhagem Celular , Criança , Subunidade alfa 1 de Fator de Ligação ao Core , Éxons , Feminino , Fibroblastos/metabolismo , Fibroblastos/patologia , Displasia Fibrosa Poliostótica/etiologia , Testa/patologia , Subunidades alfa Gs de Proteínas de Ligação ao GTP/metabolismo , Regulação da Expressão Gênica , Humanos , Dados de Sequência Molecular , Especificidade de Órgãos , Ossificação Heterotópica/congênito , RNA Mensageiro/metabolismo , Pele/metabolismo , Pele/patologia , Fatores de Transcrição/metabolismo , Transcrição Gênica
11.
J Clin Endocrinol Metab ; 58(5): 819-25, 1984 May.
Artigo em Inglês | MEDLINE | ID: mdl-6707187

RESUMO

Androgen metabolism by human epiphyseal cartilage and chondrocytes in primary culture was studied during fetal life. Testosterone (T) and androstendione (delta 4) were incubated with epiphyseal cartilage and with chondrocytes in Dulbecco's medium for 4, 12, and 24 h. Androstanedione, delta 4, dihydrotestosterone (DHT), and androstanediol were separated by bidimensional thin layer chromatography and delta 4, DHT, and T recrystallized in five different solvent systems. The results showed that delta 4 was transformed into androstanedione, DHT, and T whereas T was metabolized into delta 4 and 5 alpha-reduced androgens. In both systems (epiphyseal cartilage and chondrocytes) delta 4 was the principal metabolite produced from T. There were no major differences in the metabolic patterns of delta 4 and T between epiphyseal cartilage and chondrocytes according to sex nor gestational age (11-32 weeks). In conclusion, 5 alpha-reductase activity was found in chondrocytes from human fetuses of both sexes, but T was mainly transformed into delta 4.


Assuntos
Androgênios/metabolismo , Cartilagem Articular/metabolismo , Biotransformação , Cartilagem Articular/embriologia , Células Cultivadas , Epífises , Feminino , Feto/metabolismo , Humanos , Masculino , Técnicas de Cultura de Órgãos , Radioimunoensaio
12.
J Clin Endocrinol Metab ; 70(1): 134-40, 1990 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-2294127

RESUMO

The biological effects of dihydrotestosterone (DHT) and testosterone (T) on cultured human fetal epiphyseal chondrocytes were assessed by studying the ability of these androgens to promote DNA synthesis. DNA synthesis was evaluated by measuring [3H]thymidine incorporation into DNA. After 48-h incubation in Ham's F-12 serum-free medium, chrondrocytes were incubated with or without DHT (10(-11)-10(-8) M) or T (10(-11)-10(-8) M) in MCDB-104 serum-free medium for a further 48 h, with the addition of [3H]thymidine (5 microCi/mL) for the last 24 h. In chondrocytes from five male fetuses (12-40 weeks' gestation) DHT and T significantly stimulated DNA synthesis. The maximum stimulatory effect was obtained for DHT at 10(-10) M (P less than 0.01) and for T at 10(-6) M (P less than 0.02). In chondrocytes from four female fetuses the stimulatory effect was significant only for DHT and was maximum at 10(-10) M (P less than 0.02), whereas no effect was observed for T. Cultured chondrocytes from both male and female fetuses show the presence of proteins with high affinity and limited binding capacity (Bmax) for DHT (male fetuses: Bmax, 4.9 +/- 1.9 x 10(-15) M/mg protein; Kd, 0.43 +/- 0.24 x 10(-9) M; female fetuses: Bmax, 4.8 +/- 1.6 x 10(-15) M/mg protein; Kd, 0.63 +/- 0.19 x 10(-9) M) with no significant differences between sexes. In conclusion, our results show that androgens elicit a biological response in cultured human fetal epiphyseal chondrocytes and that DHT-binding sites are present in these cells. DHT, rather than T, seems to be the active androgen. A sex difference in the degree of androgen action is also documented.


Assuntos
Di-Hidrotestosterona/farmacologia , Desenvolvimento Embrionário e Fetal/efeitos dos fármacos , Lâmina de Crescimento/metabolismo , Receptores Androgênicos/metabolismo , Testosterona/farmacologia , Proteína de Ligação a Androgênios/metabolismo , Células Cultivadas , DNA/biossíntese , Feminino , Idade Gestacional , Lâmina de Crescimento/efeitos dos fármacos , Humanos , Masculino , Receptores Androgênicos/efeitos dos fármacos , Timidina/metabolismo
13.
J Clin Endocrinol Metab ; 75(1): 140-4, 1992 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-1619002

RESUMO

The effects of T3 on cultured human fetal epiphyseal chondrocytes were assessed by studying its effects on DNA synthesis and alkaline phosphatase activity. DNA synthesis was evaluated as follows: after 48-h incubation in Ham's F-12 serum-free medium, cultured chondrocytes were incubated with or without T3 (0.1-100 nM) in MCDB-104 serum-free medium for different periods of time (2-10 days), with the addition of [3H]thymidine (5 microCi/mL) for the last 24 h. Confluent cultured chondrocytes in 25-cm2 tissue culture flasks were incubated in Ham's F-12 serum-free medium for up to 9 days with or without T3 (0.1-100 nM); the cellular cytoplasmic fraction was obtained, and alkaline phosphatase activity was evaluated using paranitrophenylphosphate as a substrate. No significant effects of T3 (0.1-100 nM) on DNA-[3H]thymidine incorporation were observed in any experiment (n = 17) for any gestational age (12-39 weeks) or for any incubation period studied (2-10 days). However, a significant (P less than 0.025 or more) stimulatory effect of T3 (0.1-100 nM) on alkaline phosphatase activity was observed after 9 days of incubation. This effect was highest for 5 nM T3 and was present in cultured chondrocytes from human fetuses of all ages studied (13-40 weeks). Cultured human fetal epiphyseal chondrocytes from human fetuses 12-40 weeks old (n = 8) showed specific nuclear binding sites for T3. The binding capacity was 27.14 +/- 2.84 fmol/100 micrograms DNA, and the Kd was 0.66 +/- 0.14 x 0.1 nM (mean +/- SEM), with no significant differences among fetal ages. In conclusion, our results show that T3 elicits a biological response in cultured human fetal epiphyseal chondrocytes and has specific nuclear binding sites. Since alkaline phosphatase is closely related to the mineralization of epiphyseal cartilage, these results suggest that thyroid hormones could regulate this process.


Assuntos
Lâmina de Crescimento/citologia , Lâmina de Crescimento/efeitos dos fármacos , Tri-Iodotironina/farmacologia , Fosfatase Alcalina/metabolismo , Núcleo Celular/ultraestrutura , DNA/biossíntese , Feto/citologia , Feto/metabolismo , Lâmina de Crescimento/embriologia , Humanos , Receptores dos Hormônios Tireóideos/análise , Tri-Iodotironina/metabolismo
14.
J Clin Endocrinol Metab ; 84(8): 2978-81, 1999 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-10443709

RESUMO

Proliferation and differentiation of chondrocytes from growth cartilage are modulated by hormones and growth factors, among which TGF-betas have been recognized as some of the more potent regulators although their specific cell effects on cartilage physiology are not fully understood. Primary human fetal epiphyseal chondrocytes (HEFC) constitutively produce TGF-beta1 at different times of culture progression. Treatment of 48-h. serum-deprived semiconfluent HFEC with 0.1-50 ng/ml of TGF-beta1 for 48-h. decreased (3H)Thymidine incorporation by 25-50 % and cell number by 25 %. In addition, IGFBP-3, the main insulin-like bonding protein produced by HFEC, showed a slight increase by TGF-beta1 in culture media. The changes in IGFBP-3 protein levels correlated well with its mRNA, indicating that TGF-beta1 is able to up-regulate IGFBP-3 synthesis in chondrocytes. Nevertheless, the IGFBP-3 accumulation in culture media does not produce a clear growth inhibitory effect on chondrocytes. Thus, we conclude that even though TGF-beta1 is able to up-regulate IGFBP-3, the growth inhibitory action produced by TGF-beta1 is not mediated by IGFBP-3 increase and appears to be mainly a direct TGF-beta1 effect on HFEC.


Assuntos
Condrócitos/efeitos dos fármacos , Epífises/efeitos dos fármacos , Proteína 3 de Ligação a Fator de Crescimento Semelhante à Insulina/biossíntese , Fator de Crescimento Transformador beta/farmacologia , Divisão Celular/efeitos dos fármacos , Células Cultivadas , Condrócitos/fisiologia , Relação Dose-Resposta a Droga , Epífises/citologia , Epífises/metabolismo , Feminino , Feto , Humanos , Gravidez
15.
Eur J Endocrinol ; 139(3): 330-3, 1998 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-9758445

RESUMO

OBJECTIVE: Deficit of the testosterone converting enzyme 17-beta-hydroxysteroid dehydrogenase (17beta-HSD) has been shown to be responsible for male pseudohermaphroditism (MPH). We analysed the gene encoding 17beta-HSD type 3 (17beta-HSD3) in a patient with MPH. METHODS: We studied a 46, XY new-born diagnosed as having MPH. The child also had other congenital disorders, including a giant omphalocele and Fallot's tetralogy, and died of post-surgical complications at age 4.5 months. Basal hormonal levels, and after human chorionic gonadotrophin stimulation, suggested a deficiency in 17beta-HSD as the biochemical defect underlying this MPH. PCR amplification and subsequent sequencing of all coding exons of the 17beta-HSD3 gene were performed on genomic DNA from the patient and both parents. Messenger RNA was extracted from the patient's testis and 17beta-HSD3 cDNA was synthesized, PCR amplified and sequenced. RESULTS: Sequencing revealed the presence of a homozygous missense mutation (R80W) in exon 3 of the 17beta-HSD3 gene, which was also present in single doses in both parents, in accordance with the recessive inheritance of the defect. No other mutation was found, and cDNA sequencing confirmed correct synthesis and processing of 17beta-HSD3 mRNA. CONCLUSIONS: Confirming the abnormal delta4-androstenedione/testosterone ratios that suggested 17beta-HSD deficiency, a homozygous missense mutation in the gene coding for this enzyme was identified in the patient with MPH. This study adds further genetic evidence to the role of 17beta-HSD3 in male sexual development. There is no evidence supporting the association of this mutation in 17beta-HSD3 with the congenital malformations other than MPH present in the child.


Assuntos
17-Hidroxiesteroide Desidrogenases/genética , Transtornos do Desenvolvimento Sexual/genética , Mutação/genética , 17-Hidroxiesteroide Desidrogenases/deficiência , DNA/análise , DNA/genética , Transtornos do Desenvolvimento Sexual/enzimologia , Transtornos do Desenvolvimento Sexual/patologia , Eletroforese em Gel de Ágar , Éxons/genética , Humanos , Recém-Nascido , Masculino , Mutação/fisiologia , Fenótipo , Reação em Cadeia da Polimerase , RNA Mensageiro/análise , RNA Mensageiro/genética , Testículo/patologia
16.
J Steroid Biochem Mol Biol ; 53(1-6): 573-8, 1995 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-7626512

RESUMO

In view of the inconclusive data concerning the role of androgen-binding protein (ABP) in male reproductive physiology, we thought it would be pertinent to make several transgenic mouse lines overexpressing the rat ABP gene to unravel its role in Sertoli cell and epididymal homeostasis. Heterozygote transgenic mouse lines carrying the 5.5 kb ABP rat genomic DNA were produced by pronuclear microinjection. Northern blot analysis showed overexpression of rat ABP (rABP) mRNA in the testis of transgenic mice compared to rat testis control. rABP was appropriately expressed in Sertoli cells as demonstrated by in situ hybridization analysis. Sertoli cell number is increased in the seminiferous tubules of mice overexpressing rABP compared to non-transgenic littermates and scattered Sertoli cells present vacuolated-like cytoplasms, PAS and osmium negative. Compared to the wild type, the transgenic mice exhibited reduced fertility and focal damage in seminiferous epithelium characterized by morphological features compatible with programmed cell death.


Assuntos
Proteína de Ligação a Androgênios/genética , Proteína de Ligação a Androgênios/fisiologia , Animais , Feminino , Expressão Gênica , Genes , Tamanho da Ninhada de Vivíparos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos DBA , Camundongos Transgênicos , RNA Mensageiro/genética , Ratos , Reprodução , Testículo/fisiologia
17.
Clin Chim Acta ; 267(2): 167-81, 1997 Nov 28.
Artigo em Inglês | MEDLINE | ID: mdl-9469251

RESUMO

We compared two binding assays for growth hormone binding protein (GHBP) measurements, which differ in the method of bound and free GH separation: HPLC-gel filtration or dextran coated-charcoal adsorption (DCC). Two pools of sera (high and medium GHBP activity) were used for quality-control assessment. Moreover, 62 samples from 34 children and 28 adults with different nutritional status were studied. Total, between- and intra-iodination coefficients of variation (CVs) from the two methods were not different. Although percentage binding measured in the pool sera significantly differed, the concentrations assessed by Scatchard plot were comparable. Results obtained by the two methods in the 62 sera were significantly correlated (r = 0.77, P < 0.001). With both methods GHBP activity correlated with chronological age and body mass index (BMI) and differed among groups with different nutritional status. Although HPLC and DCC separation methods for GHBP measurement differ in their practicability, our study demonstrates that performance and the clinical usefulness of the two methods are comparable.


Assuntos
Proteínas de Transporte/sangue , Hormônio do Crescimento/sangue , Adolescente , Anorexia Nervosa/sangue , Carvão Vegetal , Criança , Pré-Escolar , Cromatografia em Gel , Cromatografia Líquida de Alta Pressão , Dextranos , Feminino , Transtornos do Crescimento/sangue , Humanos , Cinética , Masculino , Fenômenos Fisiológicos da Nutrição , Obesidade Mórbida/sangue , Controle de Qualidade
18.
Diabetes Res Clin Pract ; 17(3): 191-8, 1992 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-1425158

RESUMO

Our study compared the effects of an angiotensin-converting enzyme inhibitor (captopril) versus a calcium antagonist (nifedipine) on proteinuria and renal function in patients with diabetic nephropathy. A randomized follow-up study was designed. Type 2 diabetic patients, with established diabetic nephropathy (proteinuria greater than 0.5 g/24 h), were treated with nifedipine (10 patients, group A) or captopril (10 patients, group B) for 6 months. Arterial blood pressure, metabolic parameters, proteinuria and renal function were measured and compared. Mean percentage differences for glomerular filtration rate, renal plasma flow and filtration fraction between the two groups were calculated. No significant differences were observed in serum glucose, glycosylated hemoglobin (hemoglobin A1c), Na+, K+ or albumin in either group or between groups. Blood pressure decreased significantly with both treatments and mean blood pressure was significantly lower in group A compared with group B at 6 months (Mann-Whitney U-test, P = 0.03). Proteinuria was similar in both groups at randomization, but after 3 and 6 months of treatment significant reductions were observed only in the group treated with captopril (P less than 0.01). A significant decrease in filtration fraction was observed in group B with an increase in group A (Mann-Whitney U-test, P = 0.03). Multiple regression analysis identified the therapeutic agent administered as an independent variable for decrease in proteinuria. It is concluded that antihypertensive treatment with captopril, but not with nifedipine, reduced proteinuria in patients with diabetic nephropathy, although a better mean blood pressure was obtained with nifedipine.(ABSTRACT TRUNCATED AT 250 WORDS)


Assuntos
Captopril/uso terapêutico , Diabetes Mellitus Tipo 2/fisiopatologia , Glomérulos Renais/fisiologia , Nifedipino/uso terapêutico , Proteinúria/tratamento farmacológico , Glicemia/análise , Pressão Sanguínea/efeitos dos fármacos , Diabetes Mellitus Tipo 2/sangue , Diabetes Mellitus Tipo 2/epidemiologia , Feminino , Seguimentos , Hemoglobinas Glicadas/análise , Humanos , Glomérulos Renais/efeitos dos fármacos , Masculino , Pessoa de Meia-Idade , Potássio/sangue , Proteinúria/sangue , Proteinúria/epidemiologia , Análise de Regressão , Albumina Sérica/análise , Sódio/sangue
19.
J Pediatr Endocrinol Metab ; 9 Suppl 3: 313-24, 1996 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-8887176

RESUMO

Growth hormone (GH) secretion assessment in the diagnosis of short stature presents certain problems in relation to the protocols designed for it and the interpretation of results. GH measurement in serum may be accompanied by IGF-I and IGFBP-3 measurements, and in some patients by GHBP measurement. Protocols for evaluating GH response to acute stimuli or spontaneous secretion are tedious, sometimes hazardous and difficult to interpret. This is due to the wide variation in responses observed in normally-growing children, to the age-dependent changes in these parameters and, in the case of GH, to the wide variation in immunoassay results. New techniques able to measure biologically-active GH molecules circulating in blood may help to simplify diagnosis. Severe idiopathic or organic GH deficiency poses no diagnostic problems. GH secretory insufficiency may be diagnosed as partial, idiopathic, isolated GH deficiency or as neurosecretory dysfunction. Clear cut-off values for these diagnoses and the possibility of a transient reversible pathology are not well established. Analysis of large series of children with different diagnoses in whom the growth pattern, either spontaneous or under rhGH treatment, final height and GH secretion re-evaluation at the end of growth were studied will help to clarify GH secretion or action abnormalities in these patients.


Assuntos
Transtornos do Crescimento/diagnóstico , Hormônio do Crescimento Humano/sangue , Testes de Função Hipofisária , Adolescente , Estatura , Criança , Transtornos do Crescimento/sangue , Transtornos do Crescimento/etiologia , Hormônio do Crescimento Humano/deficiência , Humanos
20.
Angiology ; 39(11): 981-5, 1988 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-3177963

RESUMO

Serum total thyroxine, triiodothyronine, and thyrotropin response to thyrotropin-releasing hormone (TRH-TSH test) were measured in 126 consecutive patients admitted with atrial fibrillation: 33 patients with an acute arterial limb embolism (Group I), 31 patients with an acute embolic stroke (Group II), and 62 patients without any arterial occlusion (Group III). A blunted TRH-TSH test, suggestive of thyrotoxicosis, was found in 5 patients in Group I, 8 patients in Group II, and 2 patients in Group III. The diagnosis of hyperthyroidism was confirmed in 8 patients (by repeated TRH-TSH test and scintigraphy): 4 patients in Group I (12.1%) and 4 patients in Group II (12.9%). All of them had a nonvalvular atrial fibrillation. Thyrotoxicosis should not be recognized in 6 of them if TRH-TSH test was not performed, because peripheral hormone levels were normal. Five of these 8 patients with thyrotoxicosis had reversion to sinus rhythm after treatment with carbimazole, either spontaneously or after cardioversion. This outcome prevented prolongation of anticoagulant therapy for an indefinite time.


Assuntos
Fibrilação Atrial/complicações , Embolia/complicações , Embolia e Trombose Intracraniana/complicações , Tireotoxicose/diagnóstico , Idoso , Idoso de 80 Anos ou mais , Feminino , Humanos , Masculino , Testes de Função Tireóidea , Tireotoxicose/complicações , Tireotropina/sangue
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