Prader-Willi Syndrome and PCSK1 mutation: a novel presentation of combined syndromic and monogenic obesity.

OBJECTIVE: Prader-Willi syndrome (PWS) is a genomic imprinting disorder predominantly caused by the absence of paternally expressed imprinted genes at chromosome 15q11.2-q13. The PCSK1 gene is vital for the processing of hypothalamic POMC to ACTH and α-MSH, leading to food intake suppression and increased energy expenditure. The aim of this study was to investigate whether our PWS patient had a defect in genes involved in the hypothalamic melanocortin-4 receptor (MC4R) pathway. PATIENTS AND METHODS: A 27-year-old Greek man with PWS presented to the Adult Endocrine Clinic with morbid obesity and hyperphagia. He also had obstructive sleep apnea, growth hormone deficiency, gonadal failure and metabolic disturbances. At 6 years of age, chromosomal testing confirmed PWS with a deletion in the q11q13 region of the long arm of paternal chromosome 15. RESULTS: At the age of 27 years, further genetic testing was conducted, and next generation sequencing revealed a PCSK1_pN221D_HET mutation which was confirmed by Sanger sequencing. CONCLUSIONS: Our findings suggest that different genetic abnormalities may be present in an individual with PWS and that patients with PWS may need to be investigated for PCSK1 mutations, as the finding may potentially offer a novel treatment perspective for them.

Impaired adipocyte glucose transport regulators in morbid obesity - Possible mechanisms contributing to metabolic dysfunction.

OBJECTIVE: Obesity is characterized by hypertrophy and pathological expansion of adipocytes with impaired insulin signaling causing insulin resistance (IR) and metabolic dysfunction. We recently reported decreased expression of glucose transporter-4 (GLUT4) in cultured adipocytes from visceral and abdominal subcutaneous fat depots from patients with morbid obesity and hyperinsulinemia (MOW) and with Type 2 diabetes (MODM). Subsequently, we wanted to study the molecular mechanisms of the glucose transport regulators, p85PI3K, Rab5 and Gapex5 in morbid obesity. PATIENTS AND METHODS: Primary in vitro adipocyte cultures were developed from surgical biopsies from visceral (Visc) and abdominal (Sub) and gluteal subcutaneous (Glut) fat depots from 20 lean adults and 36 adults with morbid obesity divided into two groups: 20 with MOW and 16 MODM). mRNA and protein expression (P) of p85PI3K, Rab5 and Gapex5 were studied with RT-PCR and Western Immunoblotting (WI), respectively. RESULTS: In Sub, the P of (1) p85PI3K and Gapex5 were increased in MODM and (2) Rab5 was decreased in MOW and MODM compared to the lean. In Glut, the P of p85PI3K, Rab5 and Gapex5 showed no difference between the lean and MODM. CONCLUSIONS: In Sub of MODM (1) reduced RAB5 may possibly contribute to IR and glucose transport dysfunction, (2) increased Gapex5 may be a response to decreased Rab5 in an attempt to increase glucose transport and (3) increased p85PI3K may enhance IR mediating lipid accumulation in MODM. In Glut of MODM, though, the expression of p85PI3K, Rab5 and Gapex5 seems to be similar to that found in lean individuals.

Glucose tolerance and insulin sensitivity markers in children and adolescents with excess weight.

OBJECTIVE: Type 2 diabetes mellitus (T2DM) and obesity are alarmingly increasing in children and adolescents. Hence, predictors for early metabolic abnormalities in childhood are urgently needed. We investigated glucose tolerance in children and adolescents with obesity, markers of insulin sensitivity between males and females and the potential association between the parameters measured during an OGTT (glucose, insulin, c-peptide) and prediabetes or stages of puberty. PATIENTS AND METHODS: Glucose tolerance in 89 children and adolescents with excess weight, aged 4-19 years, from Western Greece was studied. A 3-hour OGTT was performed and fasting glucose (FG), fasting insulin (FI), 1/FI, FG/FI, Homeostatic Model Assessment of Insulin Resistance (HOMA-IR), Quantitative insulin sensitivity check index (QUICKI), ISI Matsuda index and Insulinogenic index (IGI30), were also calculated. RESULTS: No significant differences were observed in glucose values between males and females. Insulin and c-peptide concentrations were higher in the girls at several time points. FG/FI was significantly higher in the boys. Girls with obesity may be at higher risk for future insulin resistance. CONCLUSIONS: Better surveillance of pubertal girls with obesity is crucial and can be achieved using additional information provided by an OGTT, since they appear to be at a higher risk for beta-cell exhaustion. During the OGTT, not only are the baseline and 2-hour glucose and insulin measurements useful for predicting future metabolic risks and development of T2DM in children and adolescents with obesity, but additional time measurements may also be helpful.

Is parity a risk factor for excessive weight gain during pregnancy and postpartum weight retention? A systematic review and meta-analysis.

The aim of this study was to systematically review and meta-analyse the associations between parity, pre-pregnancy body mass index (BMI), gestational weight gain (GWG) and, when included, postpartum weight retention (PPWR). Papers reporting associations between parity and BMI and/or GWG in adult women were eligible: 2,195 papers were identified, and 41 longitudinal studies were included in the narrative synthesis; 17 studies were included in a meta-analysis. Findings indicated that parity was associated positively with pre-pregnancy BMI. In contrast, the role of parity in GWG was less clear; both positive and negative relationships were reported across studies. Parity was not associated directly with PPWR. This pattern of results was supported by our meta-analysis with the only significant association between parity and pre-pregnancy BMI. Overall, parity was associated with higher pre-pregnancy BMI; however, the role of parity in GWG and PPWR remains unclear, and it is likely that its influence is indirect and complex. Further research to better understand the contribution of parity to maternal obesity is warranted.

Novel application of IGF-I and IGFBP-3 generation tests in the diagnosis of growth hormone axis disturbances in children with beta-thalassaemia.

OBJECTIVE: Children with beta-thalassaemia major (beta-thal) frequently have growth retardation in the presence of low serum IGF-I and a normal GH response to pharmacological stimulation suggesting that they have GH insensitivity (GHIS). This study was carried out to study the cause of their growth retardation. DESIGN: We studied IGF-I and IGFBP-3 generation after exogenous GH administration for four days, in 15 prepubertal controls (C) and 41 prepubertal beta-thal patients divided into three groups according to their growth status: (Group 1) 15 with normal growth (N-thal) (Group 2) 16 with decelerated growth (D-thal) and (Group 3) 10 with short stature (S-thal), in order to determine whether GHIS is the cause of their growth retardation. MEASUREMENTS: IGF-I and IGFBP-3 were measured daily, before and for 4 days after daily administration of 0.1 IU/kg hGH, in 3 groups of prepubertal beta-thal patients and normal controls. RESULTS: N-thal and C had similar basal serum IGF-I (142 +/- 52 and 196 +/- 56 ng/ml, respectively) and IGFBP-3 concentrations (2.07 +/- 0.49 and 2.66 +/- 0.41 mg/l, respectively) as well as a similar percent increase of IGF-I (101 +/- 23% and 104 +/- 37%, respectively) and IGFBP-3 (52 +/- 36%, and 38 +/- 14%, respectively) during the generation tests. S-thal and D-thal had significantly lower basal IGF-I and IGFBP-3 concentrations (85 +/- 42 and 101 +/- 36 ng/ml; and 1.60 +/- 0.49 and 1.79 +/- 0.52 mg/l, respectively) as compared to N-thal and C (P < 0.001 and P < 0.005, respectively), and a significantly higher percent increase of IGF-I and IGFBP-3 during the generation tests (249 +/- 43 and 161 +/- 76%; and 121 +/- 99 and 73 +/- 35%, respectively) as compared to N-thal and C (P < 0.0001 and P < 0.01, respectively). Twenty-five percent of the growth retarded patients had classic GH deficiency (GHD) and percent increases of IGF-I and IGFBP-3 in the generation tests (164 +/- 86% and 80 +/- 49%, respectively) which were similar to those of the remaining growth-retarded children. CONCLUSION: The greater percent increases of IGF-I and IGFBP-3 in the generation tests of the growth retarded beta-thal patients, both with and without GHD, strongly suggest impaired GH secretion rather than GHIS as the cause of their growth retardation. We conclude that the IGF-I and IGFBP-3 generation tests are useful tools for the study not only of GHIS but also of GH secretory disorders in patients with beta-thal and short stature that can easily be performed in an outpatient setting as an initial test to identify the patients that may benefit from GH therapy.

Prenatal diagnosis and treatment of congenital adrenal hyperplasia and consequences in adults.

Prenatal diagnosis of congenital adrenal hyperplasia (CAH) due to 21-hydroxylase deficiency is possible using chorionic villus and amniotic fluid cells for DNA analysis of the CYP21B gene and the C4 and HLA class I and II genes. Mutations can be identified on 95% of the chromosomes using Southern blot analysis and selective amplification of the CYP21B gene by polymerase chain reaction followed by allele-specific hybridization with oligonucleotide probes for a panel of nine known CYP21B mutations. Prenatal treatment with dexamethasone at doses between 0.5 and 2 mg/day is successful in three-quarters of the treated cases in eliminating or reducing the masculinization of the affected female external genitalia. The mother must be closely monitored for any adverse effects to her health such as hypertension or glucose intolerance. Prenatal dexamethasone treatment does not seem to have an adverse effect on growth, psychological development, or school performance except for increased shyness, internalizing and decreased social behavior. Animal studies, however, have shown abnormalities in the fetal brain and behavior of the animals. Prenatal dexamethasone treatment is still offered with caution to the parents of a potentially affected CAH female.

Growth hormone (GH) provocative testing frequently does not reflect endogenous GH secretion.

GH secretion was studied in 73 children with classical GH deficiency or GH neurosecretory dysfunction (GHND), intrinsic short stature, or normal stature. The GH-deficient group was defined by a peak GH secretory response below 10 ng/ml to all provocative tests (arginine, L-dopa, insulin hypoglycemia, and clonidine). GHND was defined by a mean serum 24-h GH concentration below 3 ng/ml, with a normal response (greater than or equal to 10 ng/ml) to provocative testing. Twenty-one GH-deficient children, 21 children with GHND, and 18 short control children underwent provocative GH testing and a 24-h study with GH sampling every 20 min. A group of 13 normal stature control children also underwent 24-h GH sampling. The mean stimulated peak serum GH level [4.7 +/- 0.6 (+/- SEM) ng/ml] in the GH-deficient group was significantly below that in the GHND (19.5 +/- 1.7 ng/ml) and short control groups (24.0 +/- 3.5 ng/ml; P less than 0.01). The mean 24-h serum GH concentration was reduced in GH-deficient (1.5 +/- 0.2 ng/ml) and GHND (2.0 +/- 0.1 ng/ml) children compared to those in short (5.6 +/- 0.5 ng/ml) and normal stature (5.8 +/- 0.8 ng/ml) control children (P less than 0.01). Peak GH concentrations after provocative testing correlated poorly with 24-h mean concentrations in GH-deficient, GHND, and short control children (r = 0.38, 0.23, and 0.41, respectively; P = NS for all groups). Mean serum GH concentrations from blood sampling intervals of 12 h (day/night; 0800-2000/2000-0800 h, respectively) or even 6 h (day; 0900-1500 h) were statistically different in GHND or GH-deficient groups compared to those in control children; however, there was significantly more overlap for individual children using the 6- and 12-h daytime intervals than for the 24-h data. Plasma somatomedin-C/insulin-like growth factor I correlated with mean 24-h GH concentration endogenous secretion (r = 0.7; P less than 0.001). These data suggest that provocative GH testing frequently does not correlate with endogenous GH secretion.

Effect of a potent gonadotropin releasing hormone antagonist on pulsatile testosterone and gonadotropin secretion in the male nonhuman primate.

A potent gonadotropin releasing hormone (GnRH) antagonist [Ac-delta 3Pro1, pFDPhe2, DTrp3,6]-GnRH was given to adult male monkeys to determine the acute effect on pulsatile testosterone and gonadotropin secretion. Blood was drawn at 30 min intervals over 54 h without anesthesia using a mobile vest and tether assembly to support an indwelling catheter. After a 6 h control period, 0.1, 1.0, 2.0, 4.0 mg GnRH antagonist/kg bw in 1 ml corn oil sc, was given to intact adult male monkeys. The highest dose of GnRH antagonist decreased circulating testosterone within 6 h and for approximately 24-36 h duration. These data demonstrate that this GnRH antagonist can reduce serum testosterone both acutely and for intervals greater than 24 h and that the effective dose in intact animals is several-fold (up to 20 times) greater than in castrate animals.

Growth hormone neurosecretory dysfunction. A treatable cause of short stature.

Pulsatile growth hormone (GH) secretion was assessed in a subgroup of short children to determine whether they had GH secretory abnormalities, and these results were compared with those of normal and GH-deficient children. This subgroup of children was defined as having GH neurosecretory dysfunction and met the following criteria: height, less than first percentile; growth velocity, 4 cm/yr or less; bone age, two or more years behind chronological age, normal findings from provocative GH tests (peak, greater than or equal to 10 ng/mL), low somatomedin-C level, and abnormal 24-hour GH secretory patterns. When compared with controls, both children with GH neurosecretory dysfunction and GH-deficient patients had a significant decrease in parameters relating to the total GH secretion during the 24-hour period. As with GH-deficient children, the group with GH neurosecretory dysfunction more than doubled their growth velocity after replacement therapy with exogenous human GH during the first year of treatment. As a result of these detailed studies on pulsatile GH secretion, we suggest that there is a spectrum of GH secretory abnormalities from absolute deficiency to an intermittent irregularity in GH secretion.

Male sexual development in the nonhuman primate. III. Sertoli cell culture and age-related differences.

Homogenous preparations of primary Sertoli cell cultures were obtained from the testes of the macaque of different ages. The characteristics of Sertoli cells were confirmed by electron microscopy. Sertoli cell cultures were segregated into three developmental age groups: prepubertal, pubertal, and adult. The highest response to follicle-stimulating hormone [FSH (NIH-FSH-S13) as measured by cAMP and testosterone to estradiol conversion occurred in Sertoli cells from pubertal animals, whereas the responses were diminished in cells from both younger and older animals. Specific binding of 125I-human FSH was also increased in Sertoli cells prepared from pubertal animals when compared to cells from the other two age groups. These data demonstrate: 1) the utility of primate Sertoli cells as an in vitro model, and 2) the age-related differences in monkey Sertoli cell response to FSH and to specific FSH receptor binding.

Male sexual development in the monkey. II. Cross-sectional analysis of pulsatile hypothalamic-pituitary secretion in castrated males.

Pulsatile secretion of serum gonadotropins was studied in 16 castrated monkeys from 4 weeks of age through adult life. Animals were castrated at various ages from birth through adult life. Although some studies of the gonadotropin-secretory patterns were longitudinal in nature, most comparisons were cross-sectional. On the basis of our observations, we have arbitrarily grouped the animals into 4 developmental ages: postnatal (less than 7 months), prepubertal or juvenile (7-27 months), pubertal (28-59 months), and adult (greater than or equal to 60 months). In carrying out these studies, blood was withdrawn at 15-min intervals over 24 h without anesthesia using a mobile vest and tether assembly to support an indwelling catheter. GnRH challenge tests were done on 1 or more occasions on all animals. Plasma samples were analyzed for concentrations of FSH and LH by established RIAs and an in vitro bioassay for LH. During the frequent sampling period (24 h for all except postnatal animals), the amplitude of gonadotropin pulses was greatest in adult animals followed by postnatal and pubertal monkeys. During pubertal development, there was a marked increase in the magnitude of gonadotropin pulses, and remarkedly, there was a substantial increase in the LH bioassay: RIA (greater than 5:1) by adult life. GnRH challenge tests of gonadotropins correlated with these observations. Time series analysis was applied to the data for objective statistical characterization of cyclic patterns. Our findings can be summarized: 1) during pubertal maturation there is a change in amplitude but not frequency of gonadotropin pulses, 2) pubertal development of the hypothalamic-pituitary axis advances in the absence of gonadal feedback, and 3) there is a significant increase in the LH bioassay: RIA during pubertal development. We conclude that the castrate monkey is a valuable adjunct to direct clinical investigations of the mechanisms controlling human sexual development.

Male sexual development in the monkey. I. Cross-sectional analysis of pulsatile hypothalamic-pituitary-testicular function.

Pulsatile secretion of serum gonadotropins and testosterone was studied in 46 monkeys of varying ages from 9 days of age through adult life. Although some of the hormonal analysis was longitudinal in nature, most comparisons were cross-sectional. On the basis of pulsatile secretory patterns, hCG and GnRH stimulation, skeletal age, testicular volume, and histology, we have arbitrarily defined four developmental age groups: postnatal (less than 7 months), prepubertal or juvenile (7-27 months), pubertal (28-59 months), and adult (greater than or equal to 60 months). In accomplishing the pulsatile studies, blood was withdrawn at 15-min intervals over 24 h without anesthesia using a mobile vest and tether assembly to support an indwelling cannula. GnRH and hCG challenge tests were done on one or more occasions on all animals. Plasma samples were analyzed for concentrations of FSH, LH, testosterone, dihydrotestosterone and delta 4-androstenedione by established RIAs and an in vitro bioassay for LH. During the frequent sampling period of 24-h duration for all except postnatal animals, testosterone pulses of large amplitude (up to 8-fold) occurred in postnatal, pubertal, and adult animals. Pulsatile gonadotropin secretion was seen at all ages; however, the highest pulses (up to 15-fold) occurred in prepubertal animals even though this was an infrequent occurrence. Time series analysis techniques were applied for objective statistical characterization of cyclic patterns. Basic rhythms corresponding to 50- to 90-min frequency cycles in gonadotropin secretion were identified. Substantive differences between LH concentrations by bioassay and RIA were seen infrequently. Our findings illustrate that: 1) circulating gonadotropin and testosterone pulses change in amplitude but not necessarily frequency during pubertal development, and 2) primate models are a useful paradym for the longitudinal study of human male sexual development. We conclude that where direct human investigation may be limited, much can be learned by study of these primate surrogates.

Effect of GnRH antagonist, [Ac-delta 3Pro1, pFDPhe2, DTrp3,6] GnRH, on pulsatile gonadotrop in secretion in the castrate male primate.

Three different doses of a potent antagonist to GnRH, [Ac-delta 3Pro1, pFDPhe2, DTrp3,6], GnRH were compared in adult male monkeys to determine the acute effect on pulsatile gonadotropin secretion. In accomplishing these studies, blood was drawn at 15 min intervals over 24-30 h without anesthesia using a mobile vest and tether assembly to support an indwelling cannula. After a 3 to 6 h control period, 2.0, 0.2 or 0.02 mg GnRH antagonist/kg bw in 1 ml corn oil sc, was given to castrate adult monkeys. The highest dose decreased circulating LH (by radioimmunoassay and bioassay) and FSH whereas the intermediate dose decreased LH only. The higher dose produced both a more prolonged and greater reduction in circulating gonadotropins. These data demonstrate that this GnRH antagonist can reduce serum gonadotropins both acutely and for intervals greater than 24 h.