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Study Objective: The stimulation of the follicle-stimulating hormone receptor (FSHR) by circulating FSH or some activating mutations of the FSHR may play a causal role in the development of granulosa cell tumors of ovaries., Study Design: We evaluated four patients with ovarian juvenile granulosa cell tumors (age range, 2.4 to 7.2; median, 2.9 years) and five healthy pubertal girls (age range, 16 to 18.5; median, 16.8 years) for activating mutations in exon 10 of the FSHR. The patients were followed and evaluated clinically. Genomic DNA was extracted from the peripheral blood. Exon10 of the FSHR was evaluated for mutations., Results: All four patients presented with signs of precocious puberty. One patient, who had markedly accelerated growth velocity and advanced bone age, developed central precocious puberty after the removal of her tumor. Another patient was diagnosed to have a left ovarian cyst without tumor recurrence approximately 3.3 years after the removal of the tumor. Activating mutations were not found, but previously reported polymorphisms (Ser680Asn and Ala307Thr) of the FSHR were detected in three of four patients and in three of five controls. The follow-up period of these four patients ranged from 4.5 to 8.8 years, with a median value of 6.7 years., Conclusions: We did not find any activating mutation in exon 10 of the FSHR in our patients, and one patient developed precocious puberty after removal of her tumor. The development of ovarian tumors in these patients may have been caused by mutations at other exons of the FSHR and G protein subunits, so the association noted between central precocious puberty and granulosa cell tumors might not be coincidental.

Puberty-timing measures have historically been used as indicators of adequate nutrition and growth. More recently, these measures have been examined in relation to exposure to estrogenic or antiandrogenic agents, as well as other environmental factors. The scientific community has debated whether puberty timing is occurring earlier today than in the mid-1900s in the United States and, if so, whether environmental factors play a role; however, no one has asked a multidisciplinary panel to resolve this question. Thus, a multidisciplinary expert panel jointly sponsored by the US Environmental Protection Agency, the National Institute of Environmental Health Sciences, and Serono Symposia International was convened to examine the evidence of a secular trend, identify potential environmental factors of concern, and identify research needs regarding environmental factors and puberty timing at "The Role of Environmental Factors on the Timing and Progression of Puberty" workshop. The majority of the panelists concluded that the girls' data are sufficient to suggest a secular trend toward earlier breast development onset and menarche from 1940 to 1994 but that the boys' data are insufficient to suggest a trend during this same period. The weight-of-the-evidence evaluation of human and animal studies suggest that endocrine-disrupting chemicals, particularly the estrogen mimics and antiandrogens, and body fat are important factors associated in altered puberty timing. A change in the timing of puberty markers was considered adverse from a public health perspective. The panel recommended research areas to further our understanding of the relationships among environmental factors, puberty-timing outcomes, and other reproductive and adult disease at the individual and population levels.

Serono Symposia International convened an expert panel to review the impact of environmental influences on the regulation of pubertal onset and progression while identifying critical data gaps and future research priorities. An expert panel reviewed the literature on endocrine-disrupting chemicals, body size, and puberty. The panel concluded that available experimental animal and human data support a possible role of endocrine-disrupting chemicals and body size in relation to alterations in pubertal onset and progression in boys and girls. Critical data gaps prioritized for future research initiatives include (1) etiologic research that focus on environmentally relevant levels of endocrine-disrupting chemicals and body size in relation to normal puberty as well as its variants, (2) exposure assessment of relevant endocrine-disrupting chemicals during critical windows of human development, and (3) basic research to identify the primary signal(s) for the onset of gonadotropin-releasing hormone-dependent/central puberty and gonadotropin-releasing hormone-independent/peripheral puberty. Prospective studies of couples who are planning pregnancies or pregnant women are needed to capture the continuum of exposures at critical windows while assessing a spectrum of pubertal markers as outcomes. Coupled with comparative species studies, such research may provide insight regarding the causal ordering of events that underlie pubertal onset and progression and their role in the pathway of adult-onset disease.

Background: McCune-Albright syndrome (MAS) is characterized by a triad of gonadotropin-independent precocious puberty, café au lait skin pigmentation and fibrous dysplasia of bone. MAS is due to activating mutations of GNAS, the gene encoding Gsalpha. Interest exists in the use of GNAS mutation analysis to make a definitive diagnosis when the phenotype is not diagnostic, i.e. in partial forms of MAS. The utility of using peripheral blood for mutation analysis in this setting has not been thoroughly evaluated., Objective: We performed a systematic analysis of genomic DNA for the detection of GNAS activating mutations in girls with MAS who presented with precocious puberty to evaluate whether identification of an activating mutation in peripheral blood is related to the presence of other features of MAS., Study Design: Genomic DNA was isolated from blood from 13 girls with gonadotropin-independent precocious puberty. A polymerase chain reaction (PCR)-based technique was performed for GNAS mutation identification., Results: GNAS activating mutations were identified in 4 patients, all of whom had classic MAS based on clinical evidence., Conclusions: Detection of activating mutations in leukocyte genomic DNA extracted from peripheral blood samples from girls with gonadotropin-independent precocious puberty was associated with the presence of other phenotypic manifestations of MAS. Until improvements in the diagnostic utility of GNAS activating mutation analysis from leukocyte genomic DNA occur, such testing in patients with atypical forms of MAS should continue to be reserved for research settings.

This report describes the use of bicalutamide and anastrozole in two subjects with familial male-limited precocious puberty. Clinical improvements include decreased facial acne and pubic hair. Most importantly, a marked decrease in growth velocity and skeletal advancement has been achieved.

While ovarian cancer is a leading cause of death in females today, the molecular, genetic, and environmental factors that initiate and support the progression of this disease are still only partially understood. The extracellular signal-regulated kinase (ERK) signaling pathway is a major contributor to cellular growth, differentiation and survival. Recently, we reported that this pathway is constitutively activated in ovarian cancer cells, and that by using RNA interference (RNAi) for ERK1 and ERK2, we were able to significantly suppress the number of viable tumor cells. In the present study, we have further investigated the mechanisms by which RNAi for the ERK kinases decreased viability in these cancer cells. It was determined that treatment of the cancer cells with small inhibitory RNAs (siRNAs) directed against ERK1 and ERK2 leads to the induction of apoptosis and necrosis by four hours following treatment. Additionally, we found that primary, nonmalignant ovarian cells do not respond similarly to ERK siRNA treatment and that these cells fail to die following treatment. Data presented show that ERK2 expression is more difficult to silence, depending upon cell type being examined and that silencing ERK1 expression alone is sufficient to significantly decrease tumor cell viability.

Context: Treatment of GH-deficient adolescents in transition to adulthood remains challenging., Objective: The objective was to assess the safety and efficacy of GH in GH-deficient adolescents in transition., Patients: Fifty-eight GH-deficient adolescents (mean age, 15.8 +/- 1.8 yr; 33 males) at near completion of their linear growth participated in the study., Intervention: Baseline studies were done while subjects were on GH. Subjects were retested (insulin-induced hypoglycemia) 4 wk after GH discontinuation and reclassified as persistently GH-deficient or controls (n = 18). GH-deficient subjects were randomized to GH (n = 25, approximately 20 microg/kg.d) or placebo (n = 15)., Setting: The multicenter study was conducted over a 2-yr period., Main Outcomes: Changes in body composition, bone mineral density (BMD), quality of life (QOL), cardiovascular and metabolic markers were measured., Results: All groups had normal measures of lipid and carbohydrate metabolism, body composition, BMD, cardiac function, muscle strength, and QOL at baseline and after 2 yr. IGF-I concentrations decreased in all, but less so in the GH-group (P = 0.013). There was a greater increase in lean body mass (lesser adiposity) in the GH group than placebo at 12 months, but not at 24 months., Conclusions: 1) GH-deficient patients properly treated in childhood can have normal BMD, body composition, cardiac function, muscle strength, carbohydrate and lipid metabolism, and QOL when reaching adult height; and 2) continuation of GH therapy for 2 yr did not change these measures as compared to placebo-treated or control subjects. GH-deficient adolescents in good metabolic status at the time of epiphyseal fusion may safely discontinue GH for at least 2 yr. Follow-up is needed to determine whether GH therapy is eventually warranted in subjects treated with GH during childhood.

In March 2004 a group of 65 physicians and other health professionals representing nine countries on four continents convened in Israel to discuss the widespread public health crisis in childhood obesity. Their aim was to explore the available evidence and develop a consensus on the way forward. The process was rigorous, although time and resources did not permit the development of formal evidence-based guidelines. In the months before meeting, participants were allocated to seven groups covering prevalence, causes, risks, prevention, diagnosis, treatment, and psychology. Through electronic communication each group selected the key issues for their area, searched the literature, and developed a draft document. Over the 3-d meeting, these papers were debated and finalized by each group before presenting to the full group for further discussion and agreement. In developing a consensus statement, this international group has presented the evidence, developed recommendations, and provided a platform aimed toward future corrective action and ongoing debate in the international community.

Purpose: The use of growth hormone (GH) supplementation for intestinal adaptation among adult patients with short bowel syndrome (SBS) has provided mixed results. This report examines the effect of GH supplementation on SBS in pediatric patients., Methods: Two girls with SBS from neonatal gastrointestinal catastrophes received exogenous GH at 0.3 mg/kg per week subcutaneously and concurrent glutamine supplementation, beginning at 6 and 6(1/2) years of age. Changes in growth (height and weight) and changes in enteral and parenteral energy requirements were evaluated., Results: Treatment duration was 8 and 2.5 years, respectively. Patient weights increased from the 5th to the 41st percentile and from the 17th to the 23rd percentile, respectively. Height increased from the 1st to the 57th percentile in the former patient and increased from less than the 1st to the 17th percentile in the latter. Both patients are independent of parenteral nutrition and take enteral nutrition alone. Tolerance for enteral diets was significantly improved in each girl, with only 2 stools per day maintained in one patient., Conclusions: The data show that late exogenous treatment with GH and glutamine supplementation improved growth parameters in pediatric patients with SBS. Growth hormone and glutamine supplementation may be beneficial in promoting late intestinal adaptation in pediatric patients with SBS. These data also suggest that these adjuncts may be useful in the early phases of intestinal adaptation.

The ERK1/2 MAPK pathway is a critical signaling system that mediates ligand-stimulated signals for the induction of cell proliferation, differentiation, and cell survival. Studies have shown that this pathway is constitutively active in several human malignancies and may be involved in the pathogenesis of these tumors. In the present study we examined the ERK1/2 pathway in cell lines derived from epithelial and granulosa cell tumors, two distinct forms of ovarian cancer. We show that ERK1 and ERK2 are constitutively active and that this activation results from both MAPK kinase-dependent and independent mechanisms and is correlated with elevated BRAF expression. MAPK phosphatase 1 (MKP-1) expression, which is involved in ERK1/2 deactivation, is down-regulated in the cancer cells, thus further contributing to ERK hyperactivity in these cells. Treatment of these cancer cell lines with the proteasome inhibitor ZLLF-CHO increased MKP-1 but not MKP-2 expression and decreased ERK1/2 phosphorylation. More importantly, silencing of ERK1/2 protein expression using RNA interference led to the complete suppression of tumor cell proliferation. These results provide evidence that the ERK pathway plays a major role in ovarian cancer pathogenesis and that down-regulation of this master signaling pathway is highly effective for the inhibition of ovarian tumor growth.

Objectives: To investigate the definitive diagnosis and underlying causes of congenital hypothyroidism (CH) in eligible children through the use of a standardized protocol., Study Design: Children > or =3 years of age with CH without an identified permanent cause underwent a diagnostic algorithm. Eligible subjects had an anatomically normal thyroid or had not undergone imaging studies. After thyroxine was discontinued for 4 weeks, thyroid function tests and a thyroid ultrasound were obtained. An abnormal ultrasound was followed by a (99m)Tc thyroid scan. A perchlorate washout test was performed in subjects with a normal ultrasound but abnormal thyroid function tests. Children with normal results were followed for 1 year., Results: Of 33 children, 17 were boys. Nine (27%) had an absent or ectopic thyroid, 12 (36%) had dyshormonogenesis, and 12 (36%) had transient CH. Average thyroxine dose before medication discontinuation was 2.9 +/- 0.83 microg/kg in permanent cases versus 2.0 +/- 0.53 microg/kg in transient (P <.002). No complications from discontinuation of thyroxine occurred., Conclusions: A significant percentage of children with CH have a transient requirement for thyroid hormone. A standardized protocol with thyroid ultrasonography is a safe and sensitive approach to a trial off of thyroxine in select patients.

Objectives: To investigate the effects of growth hormone (GH) therapy on craniofacial growth and body proportions in growth hormone deficient children., Study Design: By using a cross-sectional study design, we investigated GH effects on craniofacial growth with photographic facial morphometrics, head circumference, and hand and foot size in 52 children with GH deficiency (GHD) treated with GH (0.27 mg/kg/wk) for 0.19 to 15.5 years, compared with untreated children with GHD and normal first-degree relatives. To detect disproportion and to correct for stature, age and height age (HA) SD scores were analyzed., Results: Untreated subjects with GHD had retarded facial height and width (P values=.001) compared with normal controls; small head circumference for age and HA (P=.001); small hands for age (P<.001) that were large for HA (P=.003); and small feet for age (P<.001) that were normal for HA. When compared with normal controls, GH-treated subjects had proportional facial heights but narrower facial widths. Head circumference, however, increased disproportionately to height (P=.001), becoming large for stature, and increasing with duration of therapy and cumulative GH dose (P<.001). Hands and feet grew proportionately to height., Conclusion: Growth hormone treatment with conventional doses partially corrects craniofacial deficits and does not adversely affect hand and foot growth but appears to result in excessive head circumference growth.

Granulosa cell tumors are serious ovarian neoplasms that can occur in women of all ages. While there have been numerous attempts to understand the cause of these malignancies, the pathogenesis of granulosa cell tumors (GCTs) still remains largely unknown. G-protein coupled receptor kinases (GRKs) are important regulators of signal transduction through the process of receptor desensitization and internalization. Receptors that are regulated by GRKs are members of the large family of seven-transmembrane receptors and include the follicle stimulating hormone receptor (FSHR). In granulosa cells, the FSH signaling system is responsible for cell proliferation, differentiation, and steroidogenesis. In the studies presented, we examined GRK mRNA and protein expression in nonmalignant human granulosa cells, in KGN cells, a human GCT cell line, and in a panel of human GCT samples. The KGN tumor cells express significantly less GRK4 alpha/beta protein and higher levels of GRK2 and GRK4 gamma/delta protein as compared to nonmalignant human granulosa cells. In human GCT samples, GRK4 alpha/beta protein was detected in 3 of the 13 tumor samples, whereas gamma/delta proteins expression was detected in all samples. These findings suggest that GRK protein expression is altered in GCTs and may be involved in the pathogenesis of these tumors.

Individuals with type 1 diabetes mellitus (DM1) have an increased risk of developing autoimmune thyroid dysfunction (AITD). We measured the prevalence of AITD in a pediatric DM1 population in order to examine the best combination of markers for predicting the development of AITD. A database of 1,254 patients with DM1 under 21 years of age was retrospectively screened for abnormalities in antithyroglobulin antibody (ATA), thyroid peroxidase antibody (TPO) and thyroid stimulating hormone (TSH). Charts on all 134 who had any of these serologic abnormalities were reviewed. 4.2% of the DM1 population was clinically diagnosed with AITD. Thirty-nine percent of the AITD diagnoses came within 1 year of DM1 diagnosis. Based upon evidence-based medicine statistics, TPO and TSH measurements are the most efficient and cost-effective combination of screening tests for AITD prediction and detection. The positive predictive value (of TPO and TSH) is 90%, with a positive likelihood ratio of 131.

Growth failure in children with high growth hormone (GH) levels, low insulin-like growth factor 1 (IGF-1) levels, and accelerated linear growth in response to exogenous GH is presumed to result from biologically inactive GH. A molecular diagnosis has only been made in two such patients. We analyzed the presentations and the GH-1 genes of twin Egyptian brothers with this phenotype. At 8 yr of age, the boys' heights were -4 SD. Their IGF-1 levels were 64 and 60 ng/mL, baseline GH levels were 2.1 and 11.7 mU/L, and growth hormone binding protein levels were normal. Twin B attained a peak GH level of 30.6 mU/L after L-dopa stimulation (Twin A was not tested). After 1 yr of exogenous GH, their growth velocities were >11 cm/year (>97%). Analysis of their GH-1 exons and introns revealed no mutations, but five polymorphisms were identified that have not been previously reported. The GH-1 DNA sequence was transfected into human cells and the resulting GH-1 transcripts were analyzed. Wildtype GH-1 mRNAs were observed, demonstrating that the polymorphisms do not affect transcript processing. Therefore, although no evidence of GH-1 gene mutations or abnormal GH-1 mRNA processing was found, the subjects' excellent response to exogenous GH supports a trial of GH in children with severe short stature, low IGF-1 levels and normal GH responses to stimulation testing.

Objective: We undertook a 1-year multicenter trial of tamoxifen treatment for precocious puberty in girls with McCune-Albright syndrome (MAS)., Study Design: Girls < or =10 years with classic or atypical MAS were recruited. Pretreatment history was collected for 6 months. Patients received 20 mg tamoxifen daily. Diaries were used to record bleeding. Evaluations included physical examination, bone age, pelvic ultrasound, hormone levels, and safety assessments., Results: A total of 28 girls (2.9-10.9 years of age) were enrolled from 20 centers, of whom 25 completed 12 months of tamoxifen treatment. Compared with before the study, vaginal bleeding episodes decreased (3.42+/-3.36/year vs 1.17+/-1.41/year), growth velocity slowed (SDS 1.22+/-2.65 vs -0.59+/-3.06, P=.005), and rate of bone maturation decreased (1.21+/-0.78 vs 0.72+/-0.36, P=.02). Ovarian volumes were enlarged and asymmetric throughout the study, and uterine volumes were increased. No adverse events occurred., Conclusions: Tamoxifen treatment of precocious puberty in MAS results in a reduction of vaginal bleeding and significant improvements in growth velocity and rate of skeletal maturation.

Objective: McCune-Albright syndrome (MAS) is characterized by a clinical triad of endocrinopathies, café au lait pigmentation, and polyostotic fibrous dysplasia of bone. We hypothesized that children diagnosed with fibrous dysplasia are not routinely being evaluated for coexisting endocrine dysfunction or MAS. Our objective was to prospectively screen subjects with fibrous dysplasia for endocrine disease and G(s)alpha gene (GNAS1 )-activating mutations., Study Design: Nine subjects who presented with fibrous dysplasia and were followed in orthopedic clinics were evaluated for other manifestations of MAS. Genomic DNA was isolated from blood, and mutation analysis of GNAS1 was performed., Results: On physical examination, 5 of 9 subjects were found to have café au lait pigmentation. Three of 9 subjects had TSH levels below the normal range. One of these subjects was found to have hyperthyroidism and was treated by total thyroidectomy. GNAS1 mutations were identified in 5 of 9 subjects with either monostotic or polyostotic fibrous dysplasia of bone., Conclusions: We conclude that a substantial proportion of children being followed for fibrous dysplasia of bone have unrecognized clinical and laboratory features of MAS. These children are at risk for endocrinopathy and should be screened accordingly.

Immunoassays are widely used to determine hormone levels. Antibodies directed against components of the immunoassay system can interfere with analyte concentration estimates. When unrecognized by clinicians, inappropriate clinical intervention may follow. The case of a young child with premature thelarche and elevated basal and stimulated luteinizing hormone (LH) levels is presented, in whom it is hypothesized that heterophile antibodies (HAs) caused interference in the LH immunoassay. LH concentrations were measured in two different assays: LH-microparticle enzyme immunoassay (MEIA) and LH-immunochemiluminometric assay (ICMA). To detect HA interference, LH level was remeasured after both preincubation with mouse serum to neutralize human anti-mouse antibodies, and treatment with a heterophile-blocking tube. The mean basal LH concentration by LH-MEIA was 7.4 mIU/mL and for LH-ICMA was 0.08 mIU/mL (normal range for age: 0.02-0.3 mIU/mL). LH concentration by MEIA was 0.08 mIU/mL after preincubation with mouse serum and 2.7 mIU/mL after preincubation with a heterophile blocking tube. In conclusion, HAs were identified in the serum of a child with premature thelarche. The presence of HAs led to spuriously elevated basal and gonadotropin-releasing hormone-stimulated LH concentrations, resulting in a diagnosis of central precocious puberty and unnecessary therapy. To avoid similar cases in the future, clinicians should consider the possibility of assay interference when the clinical picture is incongruent with the laboratory data.

Capillary electrochromatography using a specialty monolithic matrix was utilized in developing a rapid and highly efficient separation of isoflavones in biological materials. Without a preconcentration technique, it is relatively easy to reach ppm-ppb concentrations of these compounds in soy-based foods and verify them structurally using a photodiode array detector. With on-column preconcentration, we were able to measure low-ppb levels in human serum. Using blood samples from human volunteers, whose diet was supplemented by a soy-based product, the method has been validated for high-throughput screening of isoflavones in clinical studies.

Objective: To characterize the development of hypothyroidism in pediatric patients who receive a fixed dose of radioactive iodine (RAI)., Study Design: Medical records of children treated with fixed-dose RAI for Graves'disease between 1993 and 2001 were reviewed. Multiple variables including sex, age, thyroid hormone levels, thyroid-stimulating immunoglobulin titer, antithyroid medication use, and 24-hour radioiodine uptake were investigated as possible predictive factors for the development of hypothyroidism after treatment. All patients received RAI at a dose of between 13.8 and 15.6 mCi (average, 14.7 mCi; SD, 0.5)., Results: Permanent hypothyroidism developed in all 40 patients, although a second dose of RAI was required in one case. The average time to hypothyroidism was 77 days (SD, 32), with a range of 28 to 194 days; 75% of the patients were diagnosed with hypothyroidism between 40 and 90 days. RAI treatment was ineffective in an additional patient, who required subtotal thyroidectomy., Conclusions: We conclude that a fixed dose of RAI is effective therapy in nearly all pediatric patients with Graves'disease. Factors predicting the time course to hypothyroidism were not identified.

Multiple lines of evidence have implicated the growth hormone (GH) axis in the regulation of erythropoiesis. To test the hypothesis that GH deficiency is associated with hematologic abnormalities, we analyzed pretreatment hemoglobin levels in 100 children with GH deficiency. Hemoglobin levels were decreased in children with GH deficiency compared with age-corrected norms.

Activating mutations of the Gsalpha gene are responsible for McCune-Albright syndrome and have also been identified in sporadic tumors of the pituitary and thyroid. When associated with malignancy, activating Gsalpha mutations are known as gsp-oncogenes. We hypothesized that similar activating mutations might also account for some cases of premature thelarche and/ or granulosa cell tumors. Polymerase chain reaction and DNA sequencing was used to screen for activating mutations of Gsalpha genes in children with premature thelarche and in pathologic specimens from juvenile and adult granulosa cell tumors. Because these disorders involve over-activity of the FSH-signaling pathway, we also screened for activating mutations of the FSH receptor. No mutations were detected in either the Gsalpha or the FSHR fragment studied. Previously reported polymorphisms (Ser680Asn and Ala307Thr) of the FSHR were detected in 25/27 tumor samples and 9/9 premature thelarche samples. We conclude that activating mutations in previously identified mutation 'hot-spots' in the Gsalpha and FSH receptor genes are probably not a major cause of premature thelarche or granulosa cell tumors. In contrast, polymorphisms of the FSH receptor are common.

Ibutamoren mesylate (MK-0677), an orally active nonpeptide growth hormone (GH) secretagogue, stimulates GH release through a pituitary and hypothalamic receptor that is different from the GH-releasing hormone receptor. We evaluated the safety and tolerability and the GH-insulin-like growth factor (IGF) responses to two dosages of oral ibutamoren mesylate given to children with GH deficiency for 7 to 8 days. The patients, 18 prepubertal children (15 male, 3 female) with idiopathic GH deficiency, had a chronologic age of 10.6 +/- 0.8 years (mean +/- SD), bone age of 7.4 +/- 0.7 years, growth velocity < 10th percentile for age, height < 10th percentile for age, and a maximum GH response of < or = 10 microg/L to two different GH stimulation tests. The children were assigned as follows to one of three treatment groups with ibutamoren mesylate: 0.2 mg/kg per day for 7 days (days 1-7 or 8-14) and matching placebo for the alternate 7 days (groups I and II, respectively) or 0.8 mg/kg per day for 7 days (days 8-14, group III). On day 15 all patients received an 0.8-mg/kg dose of ibutamoren mesylate. Patients in groups I and II were studied first to assess safety at the low dose before advancement to the high dose. Hormonal profiles were evaluated on day -1 (baseline) and day 15, and the results were expressed as the change from baseline within each group. After administration of ibutamoren mesylate 0.8 mg/kg for 8 days (group III), the median increases (on day 15) from baseline were as follows: 3.8 microg/L (range, 0 to 34.3) for serum GH peak concentration (P = .001), 4.3 microg x h/L (range, 1.3 to 35.6) for the GH area under the concentration-time curve from time zero to 8 hours (AUC(0-8)) (P < .001), 12 microg/L (range, -4 to 116) for serum IGF-I (P = .01), and 0.4 microg/L (range, -0.9 to 1.5) for serum IGF-binding protein 3 (IGFBP-3) (P = .01). There was no change in serum prolactin, glucose, triiodothyronine, thyroxine, thyrotropin, peak serum cortisol, and insulin concentrations or 24-hour urinary free cortisol after administration of 0.8 mg/kg per day of ibutamoren mesylate for 8 days. We conclude that short-term administration of ibutamoren mesylate can increase GH, IGF-I, and IGFBP-3 levels in some children with GH deficiency. Thus this compound is applicable for testing its effect on growth velocity.

Saizen (recombinant growth hormone [GH]), 0.2 mg/(kg x wk), was given in an open-label fashion for an average of 51 mo to 27 children with presumed idiopathic GH deficiency who had withdrawn from a trial of Geref (recombinant GH-releasing hormone [GHRH] 1-29) because of inadequate height velocity (HV) (25 children), the onset of puberty (1 child), or injection site reactions (1 child). Measurements were made every 3-12 mo of a number of auxologic variables, including HV, height standard deviation score, and bone age. The children in the study showed excellent responses to Saizen. Moreover, first-year growth during Saizen therapy was inversely correlated with the GH response to provocative GHRH testing carried out 6 and 12 mo after the initiation of Geref treatment. These findings indicate that GH is effective in accelerating growth in GH-deficient children who do not show or maintain a satisfactory response to treatment with GHRH. In addition, they suggest that the initial response to GH therapy used in this way can be predicted by means of provoc-ative testing.

Posttranslational processing of the pro-growth hormone-releasing hormone (proGHRH) peptide can result in the formation of at least two peptide products: GHRH and the C-terminal peptide, GHRH-related peptide (GHRH-RP). While cyclic adenosine monophosphate transduces many of the actions of GHRH, other pathways also have been implicated in its actions. The aims of this study were to examine and characterize the activation of mitogen-activated protein kinase (MAPK) pathways by GHRH, and GHRH-RP in pituitary-derived GH3 cells, as well as the activation of the transcription factors that serve as substrates for these kinases. GHRH rapidly increased p44/p42 MAPK activity in GH3 cells in a protein kinase A-dependent and a protein kinase C-independent manner and stimulated the activation of the transcription factor Elk-1. By contrast, GHRH-RP and p75-92NH2 had no effect on p44/p42 MAPK phosphorylation in these cells. Additionally, we determined that all three peptides, GHRH, GHRH-RP, and p75-92NH2, rapidly and specifically increase phosphorylation of p38 MAPK and stimulate the activation of the nuclear factor CHOP. These are the first studies to demonstrate the activation of Elk-1 by GHRH and the activation of p38 MAPK and CHOP by GHRH, GHRH-RP, and p75-92NH2. We conclude that members of the GHRH family of peptides differentially activate multiple intracellular signaling pathways and suggest that the biologic actions of GHRH may be far more diverse than previously thought.

Objective: To investigate adult heights attained by patients with 21-hydroxylase deficiency and to perform a meta-analysis of height outcomes reported in this population., Study Design: A retrospective chart review of our patients >5 years of age (n = 65) who were followed up from 1978 to 1998 for 21-hydroxylase deficiency was conducted. Final height (FH) SD scores and target height (TH) SD scores were determined. The impact of sex, time of diagnosis, and compliance was assessed. Meta-analysis of results from 18 studies was performed; TH was available for 204 of 561 patients., Results: Mean FH SD score-TH SD score for our 65 patients was -1.03. For the meta-analysis, mean weighted FH SD score for all 561 patients was -1.37, whereas weighted mean FH SD score-TH SD score for the 204 patients for whom TH was available was -1.21. No difference in outcome was seen for males compared with females, although a statistically significant difference was seen for patients identified early versus late., Conclusions: Adult height in patients with 21-hydroxylase deficiency is often within 1 SD of TH. Early diagnosis and good compliance appear to improve the outcome. Rather than pursuing alternate therapies for congenital adrenal hyperplasia, efforts may instead be focused on early detection and improved compliance with traditional medical therapy.

The cause of posterior pituitary ectopia associated with anterior pituitary hormone deficiencies is unknown. We describe children with combined pituitary hormone deficiency (CPHD) or isolated GH deficiency. In all cases, magnetic resonance imaging examination revealed abnormal pituitary gland development featuring ectopic posterior lobe location and frequently hypoplastic anterior lobes. Embryonic development of the pituitary requires the coordinated expression of specific transcription factors. Mutations of the PIT-1 and PROP-1 transcription factors are responsible for CPHD in some patients with normally positioned posterior pituitaries. In mice, the Lhx3 LIM homeodomain transcription factor is required for both structural development and cellular differentiation of the pituitary gland. Thus, we hypothesized that mutations in one or both of the two human LHX3 isoforms are responsible for posterior pituitary ectopia associated with anterior pituitary hypopituitarism. Comprehensive molecular analysis of the LHX3 isoforms was performed to test this hypothesis. No loss of function mutations in the LHX3 gene were detected. In addition, analysis of PROP-1 did not reveal mutations that might cause this phenotype. These studies suggest that the abnormal processes leading to the development of CPHD or GH deficiency associated with posterior pituitary ectopia are not a result of aberrant LHX3 or PROP- 1 function, but may be caused by defects at other gene loci.

The growth hormone-releasing hormone (GHRH) gene produces a precursor molecule that contains GHRH and a carboxyl-terminal peptide that we have named GHRH-related peptide (GHRH-RP). This peptide, like GHRH, stimulates the expression of stem cell factor (SCF), an important reproductive and hematopoietic cytokine, in vitro and in vivo. In the present study, using primary cultures of rat Sertoli cells, we compared the time course of action and the level of SCF stimulation seen following treatment with GHRH-RP and GHRH. Additionally, we investigated the activity of a truncated peptide, p75-92NH2, whose sequence is contained within GHRH-RP. All three of these peptides were shown to stimulate the steady-state levels of SCF mRNA to a comparable degree. However, the time course of action for GHRH-RP differed markedly from that of GHRH. GHRH-RP and p75-92NH2, similar to GHRH, induce SCF expression, at least in part, via the activation of the protein kinase A/cyclic adenosine monophosphate intracellular signaling pathway.

The GH-releasing hormone (GHRH) precursor molecule contains a 30-amino acid C-terminal region that has been designated GHRH-related peptide (GHRH-RP). To begin to understand the physiological role of GHRH-RP, transgenic (Tg) mice that constituitively express this peptide were developed. To generate these mice, a transgene (SS-RP) was constructed by overlap primer extension PCR. This transgene, under the control of the mouse phosphoglycerate kinase gene, selectively expresses GHRH-RP, but not GHRH. Western blot analysis confirmed that the transgene produces GHRH-RP. Animals were evaluated for the effect of excess GHRH-RP on growth, fertility, behavior, stem cell factor (SCF) expression, and hematopoiesis. Northern blot and RT-PCR were used to demonstrate ubiquitous expression of the transgene in tissues from GHRH-RP Tg animals. These tissues also had marked overexpression of SCF messenger RNA compared with controls. Tg animals had significantly increased cell cycling for granulocyte-macrophage, erythroid, and multilineage progenitor cells. Transgenic animals did not differ from control mice in their growth, fertility, or behavior. These findings demonstrate, for the first time, that in vivo the C-terminal peptide of the pro-GHRH molecule is a biologically active peptide that is capable of stimulating the expression of SCF and hematopoiesis in vivo and suggests that GHRH-RP may play a role in normal blood cell development.

The prepro-GH-releasing hormone (prepro-GHRH; 12.3 kDa) precursor, like other neuropeptide precursors, undergoes proteolytic cleavage to give rise to mature GHRH, which is the primary stimulatory regulator of pituitary GH secretion. In this study we present the first model of in vitro pro-GHRH processing. Using pulse-chase analysis, we demonstrate that at least five peptide forms in addition to GHRH are produced. The pro-GHRH (after removal of its signal peptide, 10.5 kDa) is first processed to an 8.8-kDa intermediate form that is cleaved to yield two products: the 5.2-kDa GHRH and GHRH-related peptide (GHRH-RP; 3.6 kDa). GHRH-RP is a recently described peptide derived from proteolytic processing of pro-GHRH that activates stem cell factor, a factor known to be essential for hemopoiesis, spermatogenesis, and melanocyte function. Further cleavage results in a 3.5-kDa GHRH and a 2.2-kDa product of GHRH-RP. Like GHRH, there is GHRH-RP immunostaining in hypothalamic neurons in the median eminence as detected by immunohistochemistry and immunoelectron microscopy. Based on deduced amino acid sequences of the pro-GHRH processing products, several peptides were synthesized and tested for their ability to stimulate the cAMP second messenger system. GHRH, GHRH-RP, and one of these peptides [prepro-GHRH-(75-92)-NH2] all significantly stimulated the PKA pathway. This work delineates a new model of pro-GHRH processing and demonstrates that novel peptides derived from this processing may have biological action.

The rationale underlying the use of gonadotropin-releasing hormone analogues (GnRHa) to treat patients with central precocious puberty is reviewed. GnRHa are now considered the treatment of choice for patients with central precocious puberty, but the adult heights that these patients attain often fall short of what would be expected according to their genetic potential. This has led to investigations of whether adding growth hormone to GnRHa therapy can improve adult height. The results of recent combination trials are presented and analyzed.

Treatment of progressive precocious puberty in patients with McCune-Albright syndrome (MAS) has traditionally been with aromatase inhibitors, such as testolactone. However, the use of these agents has been characterized by problems with both efficacy and compliance. We report a case of MAS in which tamoxifen proved to be a successful alternative in the treatment of progressive precocious puberty. An African-American female presented with MAS at 2-5/12 years. Frequent menses, skeletal maturation and growth acceleration prompted initiation of therapy with testolactone at 22 mg/kg/d. Over the next 13 months, the patient's puberty advanced unchecked, despite progressive increases in the dose of testolactone. At age 4 years, medication was discontinued due to treatment failure. At 4-6/12 years, bone age was 10 years, predicted adult height was 137 cm, and monthly bleeding continued. Tamoxifen was then begun on an experimental basis. In response, the patient experienced immediate cessation of menses, and had an abrupt decrease in the rates of pubertal progression and linear growth. This patient has now been maintained on tamoxifen for over three years with no apparent adverse effects. GnRH analogue therapy was begun when the onset of central precocious puberty was noted. Predicted adult height has improved to 154 cm and growth velocity and skeletal maturation remain stable. Our results suggest that tamoxifen may have a valuable role in the treatment of precocious puberty in patients with MAS and may lead to superior results compared with those achieved with aromatase inhibitors.

Objective: This study was designed to test the hypothesis that hypothalamic hamartoma causes precocious puberty through a different neuroendocrine mechanism than that of normal puberty or of idiopathic precocious puberty., Design and Patients: We compared the pattern of gonadotrophin secretion among 4 girls with precocious puberty due to hypothalamic hamartoma, 27 girls with idiopathic precocious puberty, and 14 girls with normal puberty. All subjects were breast stage 3 or 4. Blood samples were obtained every 20 min for 4 h during the day (1.000 hours to 1400 h) and night (22.00 hours to 0200 h)., Measurements: LH, FSH, and prolactin were measured in each blood sample. Girls also underwent LHRH-stimulation with measurement of LH and FSH before and after stimulation., Results: There were no significant differences in mean LH level, LH peak amplitude, or LH or FSH peak frequency during either the day or the night among the three diagnostic groups. However, the mean +/- SD LHRH-stimulated peak LH levels were greater in girls with hypothalamic hamartoma than in girls with normal puberty or with idiopathic precocious puberty (194 +/- 142 vs 85 +/- 60 or 66 +/- 54 IU/l, respectively, P < 0.05). The LHRH-stimulated peak FSH level in girls with hypothalamic hamartoma exceeded the level for the normal pubertal girls (31 +/- 19 vs 17 +/- 7 IU/l, P < 0.05), but not the level for the girls with idiopathic precocious puberty (25 + 12 IU/l). The peak LH to peak FSH ratio in the girls with hypothalamic hamartoma exceeded the ratio for the girls with idiopathic precocious puberty (7.3 +/- 3.9 vs 2.6 +/- 3.0 IU/l, P < 0.05), but not the ratio for the normal pubertal girls (5.0 + 2.9). There were no significant differences in mean prolactin level, peak amplitude or frequency, or in the ratio of mean night to mean day prolactin, among the 3 diagnostic groups., Conclusions: We conclude that spontaneous gonadotrophin and prolactin secretion are similar among girls with hypothalamic hamartoma, idiopathic precocious puberty, or normal puberty. However, the increased LHRH-stimulated peak LH in the girls with hypothalamic hamartoma suggests subtle differences in neuroendocrine regulation that may underlie their more rapid pubertal maturation.

Although treatment of girls with precocious puberty should ideally restore estradiol levels to the normal prepubertal range, treatment effectiveness has usually been monitored by gonadotropin levels because estradiol RIAs have lacked sufficient sensitivity to monitor treatment effectiveness. We hypothesized that a recently developed ultrasensitive recombinant cell bioassay for estradiol would have sufficient sensitivity to demonstrate a dose-dependent suppression of estradiol during LH-releasing hormone agonist treatment and to determine whether currently used doses are able to suppress estradiol levels to the normal prepubertal range. Twenty girls with central precocious puberty were assigned randomly to receive deslorelin for 9 months at a dose of 1, 2, or 4 micrograms/ kg.day. A significant dose-response relationship was observed, with mean +/- SD estradiol levels of 16.7 +/- 6.1, 7.9 +/- 1.6, and 6.5 +/- 0.7 pmol/L at the doses of 1, 2, and 4 micrograms/kg.day, respectively (P < 0.01). The highest dose suppressed estradiol levels to just above the 95% confidence limits for normal prepubertal girls (< 0.07-6.3 pmol/L). We conclude that the ultrasensitive bioassay for estradiol has sufficient sensitivity for monitoring the response to LH-releasing hormone agonist treatment of central precocious puberty. Additionally, the observation that the deslorelin dose of 4 micrograms/kg.day did not fully restore estradiol levels to the normal prepubertal range suggests that some girls with precocious puberty may require higher doses to receive the maximal benefit of treatment. We suggest that restoration of estradiol levels to the normal prepubertal range should be the ultimate biochemical measure of efficacy, as estradiol is the key hormone that accelerates growth rate, bone maturation rate, and breast development in girls with precocious puberty.

Growth failure is a recognized feature of the Prader-Willi syndrome (PWS). Despite evidence that hypothalamic dysfunction accompanies the syndrome, the etiology of this growth failure remains controversial because most patients with PWS are obese. In order to contribute to resolution of this controversy, we performed a retrospective analysis of 16 obese and non-obese PWS children. GH deficiency was diagnosed in 12 of the 16 subjects and occurred independently of weight status. All of the non-obese subjects were GH deficient. Of the 4 GH-sufficient children, 2 were moderately obese and 2 were morbidly obese. One of these children had clinical evidence of GH deficiency including a low IGF-1 level. Only one of the children had evidence of GH deficiency and a normal IGF-1 level, a pattern that could be attributable to obesity. We conclude that most short children with PWS have growth hormone deficiency and that this deficiency probably results from hypothalamic dysfunction.