Mario Baratta

Consensus Statement on 21-Hydroxylase Deficiency from The European Society for Paediatric Endocrinology and The Lawson Wilkins Pediatric Endocrine Society

1Writing Committee: Peter E. Clayton, Royal Manchester Children’s Hospital, Manchester, UK; Walter L. Miller*,University of California, San Francisco, Calif., USA; Sharon E. Oberfield, Columbia University, New York, N.Y., USA;E. Martin Ritzen, Karolinska Institute, Stockholm, Sweden; Wolfgang G. Sippell*, University Children’s Hospital,Kiel, Germany; Phyllis W. Speiser, New York University, New York, N.Y. (*co-chairs)

Effects of a GnRH analogue (buserelin) infused via osmotic minipumps on pituitary and ovarian activity of prepubertal heifers

Abstract This work was carried out to study the pituitary and ovarian responses of prepubertal heifers to a continuous administration of GnRH, and to evaluate the effectiveness of this treatment in stimulating ovulation. Each of eight 7–8-month-old prepubertal Holstein heifers was administered with 5 μg h −1 of a GnRH analogue (buserelin) for 12 days by means of an osmotic minipump implanted subcutaneously. They were bled every 15 min for 6–8 h on Days −1, 0 (pump insertion), 3, 6, 9, 12 (pump removal) and 16. Additional blood samplings were carried out for estradiol-17β (E2) and progesterone determination. All heifers showed an immediate and marked pituitary response to GnRH stimulation, with concentrations of LH concentrations remained higher than pretreatment values, and decreased after pump removal. E2 plasma concentrations increased to 12 pg ml −1 between Day 5 and Day 9. Undetectable concentrations of progesterone in plasma during and after the treatment indicated the lack of ovulation. Our data show that a continuous infusion with GnRH seems to be effective in stimulating a long-lasting pituitary response in prepubertal heifers, and in promoting some ovarian activity and follicular development but not in inducing ovulation.

Modulation by galanin of growth hormone and gonadotropin secretion from perifused pituitary and median eminence of prepubertal male calves.

Galanin is widely distributed in the peripheral and central nervous system and has been indicated as a putative hypothalamic-hypophysiotropic hormone. This study was performed to investigate the effects of galanin on both growth (GH) and luteinizing hormones (LH) from pituitaries of young male calves. Pituitary slices (P, 500 microm in thickness) were perfused alone or coincubated with median eminence terminals (ME) in DMEM-F12 plus BSA 0.1% and antibiotics. The perifusion chambers were kept in equilibrium for 150 min, and medium samples were collected every 10 min for 240 min and stored at -20 degrees C until the measurement of LH and GH levels. Basal GH release increased up to 60% after galanin infusion (p < 0.01 vs. baseline levels) for 60 min in P alone; in P + ME coincubation, galanin-stimulated GH secretion was further increased by up to 200%. Basal LH release in chambers with P was significantly increased (up to 25%; p < 0.05) for 70 min after galanin infusion; P + ME coincubation showed a galanin-mediated increase in LH release of up to 50%. GH and LH responsiveness to exogenous GH-releasing hormone and gonadotropin-releasing hormone was not significantly modulated by galanin in our experimental model. In conclusion, galanin is demonstrated to have a significant stimulatory role in the secretion of GH and LH, with a combined action at both the hypothalamic and pituitary levels. ......................

Effects of interleukin-1β fragment (163–171) on progesterone and estradiol-17β release by bovine granulosa cells from different size follicles

Recent studies indicate that interleukin-1β may play a role in ovarian function. To explore this possibility, we examined the effects of a fragment of interleukin-1β (163–171 aa) on progesterone and estradiol-17β production by bovine granulosa cells from small ( 8 mm) follicles; in addition we investigated the distribution of monocytes/macrophages, a possible source of interleukin-1β in the follicles, to determine a possible relationship between production of the cytokine and its activity. Bovine follicles were divided according to the size, granulosa cells were obtained by aspiration and plated at a density of 105 cells/well; then different concentrations of interleukin-1β fragment (0, 0.5, 5, 20 ng/ml) with or without bFSH were added for 24 h. Progesterone (P4) and estradiol-17β (E2) concentrations in media were evaluated by validated RIAs. Interleukin-1β (0.5 and 5 ng/ml) stimulated (P < 0.05) P4 output in both small and large follicles, while FSH-induced P4 output was reduced (P < 0.05) in large but not in small follicles by all the concentrations of peptide. FSH-induced E2 output was reduced (P < 0.05) in large, but not in small follicles. Immunohisto- and cytochemistry revealed a higher number of monocytes/macrophages in large follicles. It appears from our data that: (1) interleukin-1β fragment may modulate steroidogenesis in bovine granulosa cells depending on the stage of development of the follicle, and (2) the presence of monocytes/macrophages is related to this modulation.

Effects of Interleukin-1-Beta, Interleukin-6 and Tumor Necrosis Factor-Alpha, Alone or in Association with Hexarelin or Galanin, on Growth Hormone Gene Expression and Growth Hormone Release from Pig Pituitary Cells

Objective: We studied the effects of IL-1β, IL-6 and TNF-α on GH gene expression and secretion with or without galanin and hexarelin. Methods: Pituitary cells from adult pigs were treated with IL-1β, IL-6 or TNF-α (1, 10 and 100 ng/ml), alone or in association with galanin or hexarelin (10–8M): GH mRNA was measured by RT-PCR and GH secretion by ELISA. Results: IL-1β (1, 10 and 100 ng/ml) and IL-6 (1 and 10 ng/ml) significantly (p < 0.05) enhanced GH output. IL-1β and TNF-α (1 and 10 ng/ml) reduced (p < 0.05) the galanin-induced GH secretion and IL-6 (10 ng/ml) potentiated the effect of both GH releasers (p < 0.05). GH gene expression was increased only by IL-6 at the concentrations of 1 and 10 ng/ml, either alone or in association with both galanin and hexarelin. Conclusions: We hypothesize that cytokines may play a paracrine/autocrine role in GH regulation in the pituitary independently from the intracellular pathways of the GH secretagogues.

Identification of Goat Mammary Stem/Progenitor Cells

ABSTRACT Goat mammary gland epithelial cells have been used to establish primary and permanent cell lines, but to date, no data have been available regarding mammary stem cells (MaSCs) in this species. The detection and characterization of goat MaSCs is an important task for a better understanding of the cyclic character of mammary gland development, which will also offer the potential for manipulation of lactation yield and persistency. The objective of the present study was to demonstrate that a subpopulation of goat MaSCs resides in the goat mammary gland. Mammary tissue from lactating Saanen goats (Capra hircus) was dissociated and processed to a single-cell suspension. Using an in vitro colony-forming assay, we demonstrated that distinct colony types, which expressed specific lineage markers, arose from unipotent progenitors. Using two different growth media, we showed that the frequencies of caprine clonogenic progenitors differed according to growth conditions. Goat epithelial cells were transplanted under the kidney capsule of nonobese diabetic/severe combined immunodeficient (NOD/SCID) mice, where they formed organized, bilayered structures. Our results indicate the presence of goat MaSCs in the caprine mammary gland. To our knowledge, these data represent the first description of the tissue hierarchy of the goat mammary gland and demonstrate the regenerative potential of adult goat MaSCs.

Interleukin-1β fragment (163-171) modulates bovine granulosa cell proliferation in vitro : Dependence on size of follicle

The biological effects of IL-1 on ovarian function have been considered as an inflammatory-like reaction. The peptide fragment of IL-1 (163-171 aa), which belongs to the active site of the whole protein, has been shown to exert an immunostimulatory activity without inducing inflammatory effects. The aim of this study was to investigate whether IL 163-171, alone or associated with bFSH, modulates granulosa cell proliferation; in addition, we wanted to assess if IL 163-171 interferes with FSH binding to receptors. Bovine follicles were divided according to their size. Cell proliferation, assessed by [3H]-thymidine uptake and [125I]-FSH binding, were studied in cells treated with IL 163-171(0, 0.5, 5 and 20 ng/ml) with or without bFSH (100 ng/ml) for 24, 48 or 72 h. Basal incorporation of [3H]-thymidine into granulosa cells from small follicles was always 3-fold higher (P < 0.01) than that by cells from large follicles. IL 163-171 did not show any effect in granulosa cells from large follicles but stimulated (P < 0.01) [3H]-thymidine uptake into granulosa cells from small follicles; furthermore IL 163-171 interacted positively with bFSH (P < 0.01) after 48 and 72 h. IL 163-171 significantly reduced (P < 0.05) FSH binding in cells from small follicles after 24 h, but not after 48 and 72 h. This data demonstrates that: (1) Follicular size and cell proliferation are inversely related; (2) IL 163-171 modulates granulosa cell proliferation only in cells from small follicles; and (3) the interaction between IL 163-171 and FSH does not mainly occur at receptor level.

Effects of galanin infusion on GH secretion and GHRH-induced GH release in prepubertal male lambs

Abstract The coexistence of galanin (GAL) with some classical neurotransmitters suggests that it may exert a role as a neuroendocrine modulator. The aim of this study was to investigate the GH response to porcine galanin (p-GAL) in prepubertal male lambs. In the experiments A and B the i.v. infusion of galanin (5xa0μg/kg) for 60xa0min was carried out alone (Experiment A) or combined with an i.m. injection of GHRH (0.5xa0μg/kg) 30xa0min after infusion (Experiment B); in the Experiment C, galanin (5xa0μg/kg) was infused for 300xa0min. Galanin infusion was not effective in increasing GH baseline levels ( P xa0>xa00.08); however galanin potentiated ( P