Luz Andreone

Establishment of testicular endocrine function impairment during childhood and puberty in boys with Klinefelter syndrome.

Objective  To precisely characterize the chronology of testicular endocrine function impairment during childhood and adolescence in patients with Klinefelter syndrome.

Seminiferous tubule function in delayed-onset X-linked adrenal hypoplasia congenita associated with incomplete hypogonadotrophic hypogonadism.

Objective  X‐linked adrenal hypoplasia congenita (AHC, OMIM 300200) due to mutations in the DAX‐1 gene is frequently associated to hypogonadotrophic hypogonadism (HHG, OMIM 238320). Clinical variants with delayed‐onset have been recognized. The objective of this study is to assess Sertoli cell function throughout pubertal development in patients with childhood‐onset AHC due to stop mutations in the DAX‐1 gene.

The glycan structure in recombinant human FSH affects endocrine activity and global gene expression in human granulosa cells.

The aim of this study was to analyse the biological response to different recombinant human FSH (rhFSH) glycosylation variants on the endocrine activity and gene expression at whole-genome scale in human granulosa-like tumor cell line, KGN. The effects of differences in rhFSH sialylation and oligosaccharide complexity were determined on steroid hormone and inhibin production. A microarray approach was used to explore gene expression patterns induced by rhFSH glycosylation variants. Set enrichment analysis revealed that hormone sialylation and oligosaccharide complexity in rhFSH differentially affected the expression of genes involved in essential biological processes and molecular functions of KGN cells. The relevance of rhFSH oligosaccharide structure on steroidogenesis was confirmed assessing gene expression by real time-PCR. The results demonstrate that FSH oligosaccharide structure affects expression of genes encoding proteins, growth factors and hormones essential for granulosa cells function.

Male Central Precocious Puberty: Serum Profile of Anti-Müllerian Hormone and Inhibin B before, during, and after Treatment with GnRH Analogue

We aimed to describe the functional changes of Sertoli cells, based on the measurement of serum anti-Müllerian hormone (AMH) and inhibin B during treatment with GnRHa and after its withdrawal in boys with central precocious puberty. Six boys aged 0.8 to 5.5 yr were included. AMH was low at diagnosis in patients >1 yr but within the normal range in younger patients. AMH increased to normal prepubertal levels during treatment. After GnRHa withdrawal, AMH declined concomitantly with the rise in serum testosterone. At diagnosis, inhibin B was elevated and decreased throughout therapy, remaining in the upper normal prepubertal range. In patients with testicular volume above 4 mL AMH remained higher in spite of suppressed FSH. After treatment withdrawal, inhibin B rose towards normal pubertal levels. In conclusion, AMH did not decrease in patients <1 yr reflecting the lack of androgen receptor expression in Sertoli cells in early infancy. Serum inhibin B might result from the contribution of two sources: the mass of Sertoli cells and the stimulation exerted by FSH. Sertoli cell markers might provide additional tools for the diagnosis and treatment followup of boys with central precocious puberty.

Regulation of inhibin/activin expression in rat early antral follicles.

The aim of the present study was to determine the endocrine activity of cultured early antral follicles (EAF) isolated from prepubertal diethylstilbestrol-treated rats. The effect of steroidogenic substrates and FSH on steroid, inhibin A and B, Pro-alphaC and activin A production was evaluated. Androsterone was the predominant steroid produced by EAF. The addition of androstenedione, androstenedione+FSH and progesterone stimulated oestradiol production, whereas 25-hydroxycholesterol (25-OH-Chol) increased progesterone production. Inhibin A, B, Pro-alphaC, and activin A were produced under basal conditions. The predominance of inhibin B over inhibin A was not affected by the addition of androstenedione or progesterone. Inhibin A and activin A production was stimulated by FSH. 25-OH-Chol increased Inha, Inhba and Inhbb mRNA expression and the production of the three molecular forms of inhibins but decreased activin A production. These results show that FSH and the steroid follicular microenvironment differentially modulate the gene expression of inhibin/activin subunits, their assembly and secretion.

Effect of sialylation and complexity of FSH oligosaccharides on inhibin production by granulosa cells

Granulosa cell (GC) inhibin A and B production is regulated by FSH and gonadal factors. This gonadotrophin is released as a mixture of glycoforms, which induce different biological responses in vivo and in vitro. Our aim was to determine the effect of recombinant human FSH (rhFSH) glycosylation variants on inhibin A and B production by rat GCs. Preparative isoelectro focusing was used to isolate more acidic/sialylated (pH <4.00) and less acidic/sialylated (pH >5.00) rhFSH charge analogues. Concanavalin A was used to isolate unbound and firmly bound rhFSH glycoforms on the basis of their oligosaccharide complexity. GCs, obtained from oestrogen-primed immature rats, were cultured with either native rhFSH or its glycosylation variants. Inhibin A and B were determined using specific ELISAs. Results were expressed as mean±s.e.m. Under basal conditions, inhibin A was the predominant dimer produced (inhibin A: 673±55; inhibin B: 80±4  pg/ml). More acidic/sialylated charge analogues stimulated inhibin B production when compared to inhibin A at all doses studied; by contrast, less acidic/sialylated charge analogues stimulated inhibin A production and elicited no effect on inhibin B. Glycoforms bearing complex oligosaccharides showed a potent stimulatory effect on inhibin B when compared to inhibin A production (i.e. dose 1  ng/ml: 4.9±0.5 vs 0.9±0.1-fold stimulation, P<0.001). Glycoforms bearing hybrid-type oligosaccharides favoured inhibin A production (i.e. dose 4  ng/ml 2.9±0.1 vs 1.6±0.1-fold stimulation, P<0.05). These results show that the sialylation degree as well as the complexity of oligosaccharides present in the rhFSH molecule may be considered additional factors that differentially regulate dimeric inhibin production by rat GCs.