[Show abstract][Hide abstract] ABSTRACT: Context: Aromatase (P450arom) is the key enzyme for estrogen biosynthesis and is encoded by the CYP19A1 gene. Since 1991, several molecular CYP19A1 gene alterations associated with P450arom deficiency have been described in both sexes. Objective: To detect CYP19A1 mutations in five aromatase deficient 46,XX patients and to describe the clinical follow-up from birth to puberty. To perform haplotype analysis associated with the high frequency c.628G>A splice mutation in Argentinean patients. Design: Sequencing of the coding and flanking intronic regions of the CYP19A1 gene in all patients and parents. Haplotype analysis of patients carrying the c.628G>A mutation. Patients: Clinical and biochemical findings in five new cases and one previously reported female aromatase-deficient patient (46,XX) are described. All patients presented with ambiguous genitalia at birth. Congenital adrenal hyperplasia due to 21-hydroxylase deficiency as well as other steroidogenic defects were ruled out. Results: Phenotypic variability among the affected patients was found during follow-up. Direct sequencing of the CYP19A1 gene from genomic DNA revealed one novel mutation (c.574C>T) in two patients. In silico analysis predicted the c.574C>T mutation to be probably damaging. Four out of six nonrelated patients presented with the c.628G>A splice mutation. Haplotype analysis showed that the c.628G>A splice mutation is associated with the same haplotype in our population. Conclusions: Increased knowledge on phenotypical variability found in female aromatase-deficient patients is useful to improve the detection rate in this disorder. In our population, a genetic founder defect has probably contributed to an increase in the incidence of the c.628G>A splice mutation.
No preview · Article · Nov 2014 · Journal of Clinical Endocrinology & Metabolism
[Show abstract][Hide abstract] ABSTRACT: BackgroundIGF1R gene mutations have been associated with varying degrees of intrauterine and postnatal growth retardation, and microcephaly.Objectiveto identify and characterize IGF1R gene variations in a cohort of 28 Argentinean children suspected of having IGF-1 insensitivity, who were selected on the basis of the association of pre/postnatal growth failure and microcephaly.Methods
The coding sequence and flanking intronic regions of IGF1R gene were amplified and directly sequenced. Functional characterization was performed by two in vitro assays: 1) [Methyl-3H] thymidine incorporation into DNA in fibroblast cell primary cultures from patients and controls treated with IGF-1 for 16-24 hs. 2) PI3K/Akt pathway was evaluated with phospho Akt (Ser473) STAR ELISA Kit (Millipore) in fibroblast cultures from patients and controls stimulated with IGF-1 for 10 minutes. Prepubertal clinical and GH-IGF-1 axis evaluation was followed up.ResultsWe identified three novel heterozygous missense mutations in three unrelated patients, de novo p.Arg1256Ser, de novo p.Asn359Tyr and p.Tyr865Cys. In control cells, proliferation assay showed that IGF-1significantly induced DNA synthesis at 20hs, and Akt phosphorylation assay that it significantly stimulated phosphorylation after 10 minutes (p<0.05 by ANOVA and Bonferroni Tests). However, no significant increase was observed in any of the three patient fibroblasts in both functional studies. GH therapy growth response in two patients was inconsistent.Conclusion
These variations led to failure of the IGF1R function causing pre and postnatal growth retardation and microcephaly. Microcephaly should be considered in the evaluation of SGA patients, since it seems to favor the frequency of detection of IGF1R mutations.This article is protected by copyright. All rights reserved.
No preview · Article · Jul 2014 · Clinical Endocrinology
[Show abstract][Hide abstract] ABSTRACT: StAR facilitates cholesterol entry into the mitochondria as part of the transduceosome complex. Recessive mutations in the gen STAR cause classic and nonclassic congenital lipoid adrenal hyperplasia. The aim of the study was to analyze the molecular consequences of a novel heterozygous STAR mutation in a 46,XY patient with ambiguous genitalia and adrenal insufficiency. We found a de novo heterozygous IVS-2A>G STAR mutation and the reported heterozygous p.G146A SF1 polymorphism with normal CYP11A1, FDXR, FDX1, VDAC1 and TSPO genes. RT-PCR and sequencing from patient's testicular RNA showed a -exon2 transcript and the wild-type (WT) transcript. Both 37 kDa precursor and 30 kDa mature protein were detected in COS-7 cell transfected with mutant and WT plasmids. Immunofluorescence showed almost no co-localization of mitochondria and mutant protein (delta22-59StAR). Delta22-59StAR activity was 65±13% of WT. Cotransfection with WT and delta22-59StAR plasmids reduced WT activity by 62.0% ± 13.9. Novel splice-junction heterozygous STAR mutation (IVS-2A>G) resulted in the in-frame loss of amino acids 22 to 59 in the N-terminal mitochondrial targeting signal. A misfolded p.G22_L59delStAR might interfere with WT StAR activity by blocking the transduceosome complex, causing an autosomal dominant form of StAR deficiency, explaining the clinical phenotype.
[Show abstract][Hide abstract] ABSTRACT: Context:Steroid acute regulatory (StAR) protein is a mitochondria-targeted protein that is part of the transduceosome complex crucial for transport of cholesterol to mitochondria. Recessive mutations cause classic and nonclassic congenital lipoid adrenal hyperplasia.Objective:The aim of this study was to report the clinical, hormonal, genetic, and functional data of a novel heterozygous mutation in the StAR gene found in a 46,XY patient with ambiguous genitalia and neonatal severe steroidogenic deficiency.Patient:Undetectable serum steroids with high ACTH and plasma renin activity but normal acute GnRH response were found in infancy. After gonadectomy (at 3 yr of age), serum LH and testosterone were undetectable, whereas FSH was normal but increased slowly afterward. Estrogen replacement therapy, started at 10.2 yr of age, suppressed gonadotropins (for 2 yr). However, after 1 month off estrogens, the patient showed castrated levels. At 11.9 yr old, after fludrocortisone withdrawal because of hypertension, plasma renin activity and aldosterone remained normal, suggesting mineralocorticoid recovery by a StAR-independent mechanism.Results:We found a de novo heterozygous IVS-2A>G StAR mutation and the reported heterozygous p.G146A SF1 polymorphism with normal CYP11A1, FDXR, FDX1, VDAC1, and TSPO genes. The mutant StAR transcript lacked exon 2, resulting in the in-frame loss of amino acids 22 to 59 in the N-terminal mitochondrial targeting signal. In vitro, the mutant protein exhibited reduced StAR activity in a dominant-negative manner and almost no mitochondria localization.Conclusions:A misfolded p.G22_L59del StAR might interfere with wild-type StAR activity by blocking the transduceosome complex, causing an autosomal dominant form of StAR deficiency, explaining the clinical phenotype. We speculated that estrogen might have modulated mineralocorticoid function and pubertal maturation in a human natural model lacking endogenous steroid production.
No preview · Article · Nov 2012 · The Journal of Clinical Endocrinology and Metabolism
[Show abstract][Hide abstract] ABSTRACT: In humans, steroidogenic factor 1 (NR5A1/SF-1) mutations have been reported to cause gonadal dysgenesis, with or without adrenal failure, in both 46,XY and 46,XX individuals. We have previously reported extreme within-family variability in affected 46,XY patients. Even though low ovarian reserve with preserved fertility has been reported in females harboring NR5A1 gene mutations, fertility has only been observed in one reported case in affected 46,XY individuals. A kindred with multiple affected members presenting gonadal dysgenesis was studied. Four 46,XY individuals presented severe hypospadias at birth, one of them associated with micropenis and cryptorchidism. The other 3 developed spontaneous male puberty, and 1 has fathered 5 children. Four 46,XX patients presented premature ovarian failure (one of them was not available for the study) or high follicle-stimulating hormone levels. Mutational analysis of the NR5A1 gene revealed a novel heterozygous mutation, c.938G→A, predicted to cause a p.Arg313Hys amino acid change. A highly conserved amino acid of the ligand-binding domain of the mature protein is affected, predicting abnormal protein function. We confirm that preserved fertility can be observed in patients with a 46,XY disorder of sex development due to heterozygous mutations in the NR5A1 gene.
Full-text · Article · Aug 2012 · Hormone Research in Paediatrics
[Show abstract][Hide abstract] ABSTRACT: Growth retardation is a considerable clinical problem in children with chronic kidney disease (CKD). Optimization of metabolic and nutritional parameters does not always lead to improved growth. Recombinant human growth hormone (rhGH) treatment has been used to improve height. Several studies in the literature have shown increased growth velocity, although data on the final height (FH) reached are scarce.
We assessed the effect of rhGH on FH standard deviation score (SDS) in children with CKD following renal transplantation (RTx), comparing it with patients who did not receive rhGH (control group) but were treated with the same protocol and followed up in a single Center.
Thirty-three patients received rhGH treatment until FH. Fourteen who refused rhGH therapy were included in the controls. Prognostic factors for FH and changes in glomerular filtration rate (GFR) during follow-up were also analyzed
FH SDS in rhGH-treated patients was significantly higher than in controls (-1.88 ± 1.14 vs -3.48 ± 1.19 SDS, respectively, p <0.05). In both groups, a similar reduction in GFR was observed. Height (SDS) at onset of rhGH treatment was the only statistically significant variable useful to predict response to treatment (p = 0.001).
Our findings confirm that rhGH is effective to improve FH in CKD RTx patients, without affecting kidney function.
No preview · Article · Jan 2012 · Pediatric Nephrology
[Show abstract][Hide abstract] ABSTRACT: To report genotype-phenotype correlation in a large cohort of patients.
Study of the CYP21A2 gene in 866 unrelated chromosomes of 21-hydroxylase deficiency in Argentinean patients with classic and nonclassic (NC) forms of congenital adrenal hyperplasia (CAH).
Eleven most common mutations were analysed by allele-specific polymerase chain reaction, restriction fragment length polymorphism (RFLP) or southern blot analysis. Gene sequencing was performed when no mutation was detected in one allele or the genotype-phenotype correlation was lacking.
The 11-most-common-mutation screening allowed for the detection of 88·1% of affected alleles (80·3% in the NC and 95·2% in the classic forms). p.V281L, IVS2-13A/C>G (In2) and gene deletions and large gene conversions were the most prevalent mutations. In2 (35·2%) in salt wasting (SW), p.I172N (37·3%) in simple virilizing and p.V281L (54·1%) in NC CAH were the most prevalent mutations within the clinical forms. In 7/15 p.P30L mutation alleles, a chimeric CYP21A1P/CYP21A2 gene [PromCYP21A1P; p.P30L] was detected, while 6/15 represented a single-nucleotide substitution, and in 2/15 linkage with mutations, p.[P30L; V281L] and [p.P30L; IVS2-13A/C > G; p.Q318X] was found. In two SW patients, a novel nonsense mutation, p.Q41X, was observed. In three p.V281L mutation patients, the phenotype was more severe than predicted by genotype. Sequence analysis revealed an intronic alteration in the allele carrying the p.V281L mutation [IVS2 + 5G > A; p.V281L]. An aberrant splicing in this p.V281L mutated allele explains the clinical phenotype.
A high percentage of CYP21A2 affected alleles is detected by the 11-mutation screening study. Genotype-phenotype correlation was high, but when the phenotype is more severe than predicted by genotype, presence of two alterations in one allele should be ruled out.
Full-text · Article · May 2011 · Clinical Endocrinology
[Show abstract][Hide abstract] ABSTRACT: Three novel heterozygous SF-1 gene mutations affecting multiple members of two unrelated families with a history of 46,XY disorders of sex development (DSD) and 46,XX ovarian insufficiency are described.
clinical and mutational analysis of the SF-1 gene in 9 subjects of two families.
family 1 had 2 affected 46,XY DSD subjects. One, born with severe perineal hypospadias, was raised as a male, and presented normal adolescence. The other, born with ambiguous genitalia, uterus, and mild testicular dysgenesis, was raised as a female. A W279X heterozygous mutation and an intronic deletion (g3314-3317delTCTC (IVS 4 + 8) was found in the SF-1 gene. In family 2, 4/6 affected siblings had 46,XY DSD or hypospadias. An affected 46,XX sister had normal sexual development but increased FSH levels. The 37-year-old affected mother had entered menopause. An Y183X heterozygous mutation was detected.
an extreme within-family phenotypic variability, ranging from severe prenatal undervirilization to normal pubertal development, was observed in 46,XY-affected siblings, indicating that other unknown factors might be involved in the phenotype. Low ovarian reserve and preserved fertility in 46,XX subjects can be observed in heterozygous SF-1 gene mutations.
Full-text · Article · Jan 2011 · Hormone Research in Paediatrics
[Show abstract][Hide abstract] ABSTRACT: GH deficiency (GHD) and spine irradiation (SI) have been implicated in the mechanism of reduced adult height (AH) in childhood survivors of medulloblastoma. However, growth dynamics after tumor diagnosis and the effectiveness of rhGH on AH in comparison with rhGH-untreated survivors have not been reported.
To follow height (H) SDS (HSDS) since tumor diagnosis and the effect of rhGH in GHD patients, comparing with GH-untreated GHD patients.
14 patients received rhGH treatment until AH (medulloblastoma GH-treated group, MGHGr). 19 patients refused rhGH therapy (GH-untreated control medulloblastoma group, MCGr). Standing H and sitting H (SitH) were measured.
In MGHGr, mean +/- SD HSDS decreased from 0.09 +/- 0.63 at tumor diagnosis to -1.38 +/- 0.91 at diagnosis of GHD, and to -1.90 +/- 0.72 at the onset of rhGH, p < 0.01, but it remained unchanged during rhGH (AH -2.12 +/- 0.55). MCGr HSDS (-0.25 +/- 0.88) was not different from MGHGr at tumor diagnosis, but it was -3.40 +/- 0.88 at AH, significantly lower than in MGHGr, p = 0.001. SitH SDS at AH (-4.56 +/- 0.82) was significantly lower than at the onset of rhGH (-2.86 +/- 0.75), p = 0.003, and it was not different from MCGr (-4.85 +/- 1.77).
rhGH treatment improves AH in GH-deficient childhood medulloblastoma survivors but not spinal growth.
No preview · Article · Mar 2010 · Hormone Research in Paediatrics
[Show abstract][Hide abstract] ABSTRACT: Adrenarche occurs only in higher primates, typically at 6-8 years of age in humans, when the innermost layer of the adrenal cortex, the zona reticularis (ZR), develops. This is an event of postnatal sexual maturation in which there is an increase in the secretion of adrenal androgens, mainly dehydroepiandrosterone (DHEA) and DHEA sulfate (DHEAS), not accompanied by an increase in cortisol secretion. Although adrenarche might be a multifactorial event, the main process regulating the production of adrenal androgens continues to represent one of the most intriguing mysteries in adrenal functional differentiation. Premature adrenarche, the early increase in adrenal androgen production, might be associated with ovarian hyperandrogenism and polycystic ovarian syndrome (PCOS) in females, as well as with insulin resistance in the two sexes, during puberty development and adult life, and even in childhood. However, many questions remain with incomplete answers, such as (i) the mechanisms of adrenarche and premature adrenarche, (ii) the physiological actions of adrenal androgens, acting as either direct ligands or pro-hormones, and (iii) the relationship between the activation of adrenal androgen secretion, growth restriction during fetal life, and chronic diseases in adulthood, transforming adrenal androgens in markers of diseases important for human health. Future research might contribute to provide responses for a better understanding of these questions.
No preview · Article · Jan 2010 · Current Clinical Neurology
[Show abstract][Hide abstract] ABSTRACT: Serum IGF-I and IGFBP-3 assays are used to monitor rhGH treatment. Some discrepancies in results obtained by means of different assays have been reported. The aim of this study was to establish normal ranges for circulating IGF-I and IGFBP-3 in children and adolescents of Hispanic and Italian origin. Circulating levels of IGF-I and IGFBP-3 were measured in 169 Hispanic and Italian prepubertal children and 66 adolescents of both sexes, using a chemiluminescent assay. Serum levels of IGF-I and IGFBP-3 increased from early childhood into adolescence. After pubertal peaks of IGF-I and IGFBP-3, slight decreases were observed with increasing age. Furthermore, serum IGF-I levels were significantly higher in girls than in boys, suggesting a sexual dimorphism in serum IGF-I values in late prepuberty and early puberty. Differences in IGF-I and IGFBP-3 absolute values between our study and previous studies suggest the need to establish reference ranges for each ethnic group.
No preview · Article · Dec 2009 · Journal of pediatric endocrinology & metabolism: JPEM
[Show abstract][Hide abstract] ABSTRACT: Insulin resistance (IR), abnormal lipid profile, and other features of the metabolic syndrome have been described in CYP19 gene knockout mice and in aromatase-deficient adult men but not in prepubertal affected girls.
To study insulin sensitivity, as well as the effects of estrogen, metformin and GnRHa treatment on glucose homeostasis, in an aromatase-deficient girl.
Clinical, metabolic and hormonal follow-up data, from 8 to 12 years of age, is presented.
At 9 years of age, IR (HOMA 5.6) and glucose intolerance was detected, along with high serum testosterone (2.28 nmol/l), androstenedione (4.92 nmol/l) and FSH (13.4 mIU/ml) levels. Estrogen replacement was ineffective to suppress gonadotropin and androgen levels, as well as IR. Under metformin therapy, she developed type 2 diabetes and acanthosis nigricans. GnRHa administration for 1 year resulted in marked decreases in gonadotropin and serum androgens, but severe IR persisted.
Postnatal estrogen replacement and a marked decrease of endogenous androgens failed to improve IR and glucose tolerance. We propose that, in females, the increment of androgens and/or lack of estrogens during fetal life might alter the mechanism of fetal programming of insulin sensitivity.
No preview · Article · Oct 2009 · Hormone Research
[Show abstract][Hide abstract] ABSTRACT: INTRODUCTION: cP450aromatase deficiency provides clues for the understanding of the role of aromatase in prepubertal and pubertal human health and disease. Placental aromatization of androgens protects the female fetus against the virilizing action of fetal androgens. After birth, the dual effect of aromatase deficiency, excessive androgens, and insufficient estrogens is responsible for a variable clinical picture. Nineteen cases of aromatase gene (CYP19) deficiency have been reported. PHENOTYPE: Phenotype is dependent on sex and age. In newborns, aromatase deficiency should be considered in the etiology of 46,XX DSD, after ruling out congenital adrenal hyperplasia. In prepubertal aromatase deficient girls, high levels of ovarian androgens and gonadotropins facilitate the formation of ovarian cysts. Bone mineralization can be affected and bone aging is delayed. In pubertal girls, there is poor sexual development and abnormal virilization. The phenotype may be variable according to enzyme activity level. Insulin sensitivity may be abnormal in both men and women. Finally, aromatase might also play a role in the regulation of testicular cell mass in the newborn testis. CONCLUSION: Adequate interpretation of clinical data should lead to the analysis of the CYP19 gene for diagnostic confirmation and implementation of appropriate management.
No preview · Article · Oct 2009 · Hormone Research
[Show abstract][Hide abstract] ABSTRACT: Sex hormones are modulators of the GH/ IGF-I system. We have hypothesized that the inhibition of DHEAS in treated girls with congenital adrenal hyperplasia (CAH) might affect this modulation. We analyzed serum IGF-I, IGFBP-3 and DHEAS in 17 prepubertal (Pp) and 32 pubertal (Pu) girls with CAH, under hydrocortisone replacement therapy, in the presence of normal (Gr1) or high (Gr2) serum testosterone (T) and androstenedione (A) levels. All groups had appropriate normal controls. Serum DHEAS in patients with CAH was significantly lower than in the respective controls (p < 0.04), except for Pp CAH Gr2. Serum IGF-I, but not serum IGFBP-3, in CAH subgroups was significantly higher than in the respective controls (p < 0.05), except for Pp CAH Gr2. It is concluded that glucocorticoid treatment of girls with CAH results in hypofunction of the adrenal zona reticularis. Low levels of serum DHEAS could be involved in the regulation of IGF-I biological response in target tissues. Additional studies are necessary to confirm these findings.
No preview · Article · Mar 2009 · Journal of pediatric endocrinology & metabolism: JPEM