Cristina Esteban

The exon 3-deleted/full-length Growth Hormone Receptor Polymorphism Does Not Influence the Effect of Puberty or Growth Hormone Therapy on Glucose Homeostasis in Short Non-Growth Hormone-Deficient Small-for-Gestational-Age Children: Results from a Two-Year Controlled Prospective Study

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).

Androgen binding protein is tissue-specifically expressed and biologically active in transgenic mice

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.

Human growth hormone (GH1) gene polymorphism map in a normal‐statured adult population

Objective  GH1 gene presents a complex map of single nucleotide polymorphisms (SNPs) in the entire promoter, coding and noncoding regions. The aim of the study was to establish the complete map of GH1 gene SNPs in our control normal population and to analyse its association with adult height.

Clinical, Biochemical and Morphologic Diagnostic Markers in an Infant Male Pseudohermaphrodite Patient with Compound Heterozygous Mutations (G115D/R246W) in SRD5A2 Gene

A patient with male pseudohermaphroditism and clinical diagnosis of partial androgen insensitivity in the neonatal period was studied at pubertal age for a molecular diagnosis. Hormone studies were conducted at baseline and under hCG stimulation for testosterone and dihydrotestosterone determinations at 2 months of age. Gonadectomy was performed at 4 months. At the age of 13 years genital skin fibroblasts were studied for androgen binding and 5α-reductase activity and peripheral blood DNA was available for androgen receptor (AR) and 5α-reductase (SRD5A2) gene analysis. Exons 1–8 of AR gene and exons 1–5 of SRD5A2 gene were sequenced. AR gene coding sequences were normal. SRD5A2 gene analysis revealed two heterozygote mutations (G115D and R246W), with the mother carrying the G115D and the father the R246W mutations. The compound heterozygote mutations in SRD5A2 gene explained an extremely low 5α-reductase enzyme activity in genital skin fibroblasts. Revision of hormonal data from the neonatal period revealed an increased testosterone-to-dihydrotestosterone ratio at the end of an hCG stimulation test, which concurred with the molecular diagnosis. Testis morphology at 4 months of age was normal. Clinical and biochemical differential diagnosis between partial androgen insensitivity syndrome and 5α-reductase enzyme deficiency is difficult in the neonatal period and before puberty. Our results show that in our patient the testosterone-to-dihydrotestosterone ratio would have adequately orientated the diagnosis. The two mutations in SRD5A2 gene have been described in patients of different lineages, though not in combination to date. Testis morphology showed that, during early infancy, the 5α-reductase deficiency may not have affected interstitial or tubular development.

Sertoli cell-specific expression of rat androgen-binding protein in transgenic mice: effects on somatic cell lineages

The Sertoli cells of many species produce an androgen binding protein (ABP) which carries testicular androgens to the epididymis and is thought to play a role in sperm maturation. In the present report we analyzed the morphological modifications present in Leydig, Sertoli, and peritubular cells of the testis of young adult male mice transgenic for ABP gene, which overproduce ABP in testis. By in situ hybridization we demonstrated that ABP is specifically produced by Sertoli cells. Using light and electron microscopy, we detected scattered alterations of the seminiferous tubule cells which include cell degeneration and vacuolization. Leydig and Sertoli cells present morphological signs of hyperfunctioning compensatory mechanisms which include increased amounts of lipid droplets probably due to the existence of a stimulated steroid synthesis that in turn could be a consequence of the decreased unbound testosterone and/or a direct paracrine effect of ABP. Peritubular cells also present numerous signs of hyperstimulation.

Vitamin D Stimulates Growth Hormone-Insulin-Like Growth Factor (GH-IGF) Gene Axis Expression and Potentiates GH Effect to Reverse the Inhibition Produced by Glucocorticoids in Human Growth Plate Chondrocytes

Chondrocytes from human fetal epiphyseal growth plates were obtained at first passage, serum-deprived for 48 h and incubated for a further 48 h with Dx (10 –9 to 10 –6 M ), VitD (10 –11 to 10 –6 M ) or GH (500 ng/ml) or combinations of Dx, VitD and GH. Cell proliferation was determined as 3 H-thymidine incorporation into DNA (n = 25), and gene expression of growth factors and binding proteins (IGF-I, IGF-II, IGF binding protein-3 [IGFBP3]), receptors (IGF-IR, GHR), SOX9 transcription factor and matrix proteins (aggrecan, COL2A1, COMP) were analyzed by total RNA extraction and real-time quantitative polymerase chain reaction (n = 8).

SRD5A2 gene Q126R exon 2 point mutation in unrelated Spanish male pseudohermaphrodite patients.

Three unrelated Spanish patients born with ambiguous genitalia, 46,XY karyotype, normal testosterone (T) secretion, and extremely low 5a-reductase activity in genital skin fibroblasts or absence of dihydrotestosterone (DHT) stimulation under human chorionic gonadotropin (hCG), were studied for a molecular diagnosis. Exons 1 to 8 of the AR (androgen receptor) gene and exons 1 to 5 of the SRD5A2 (steroid-5-alpha-reductase type 2) gene were sequenced in peripheral blood DNA. The AR gene coding sequences were normal. SRD5A2 analysis revealed the same exon 2 missense point mutation (Q126R) in all three unrelated patients. One was homozygous, another was a compound heterozygote (associated with an exon 4, N193S, point mutation), and the third heterozygous with no other sequence change in coding and intronic flanking regions of SRD5A2 gene. The heterozygous patient was the one presenting a milder phenotype with male gender assignment. The low 5a-reductase activity in genital skin fibroblasts in the first two patients and the increase in T/DHT ratio under hCG stimulation in the third were in accordance with the molecular diagnosis. The two mutations, Q126R and N193S, have been described in patients of different geographic and ethnic origins and could thus be considered recurrent mutations in SRD5A2 gene.