Vascular endothelial growth factor (VEGF) levels in short, GH treated children: a distinct pattern of VEGF-C in Noonan syndrome.

CONTEXT: Noonan syndrome (NS) is characterized by short stature and elevated risk of lymphedema. The mechanism underlying lymphedema may be mediated by vascular endothelial growth factors (VEGFs). OBJECTIVE: To assess the effect of growth hormone (GH) treatment on plasma insulin-like growth factor (IGF)-1, VEGF-A and VEGF-C levels in patients with NS as compared to short GH-sufficient children. DESIGN: Retrospective, comparative. SETTING: Endocrinology department of a tertiary pediatric medical center. PATIENTS AND METHODS: Plasma IGF-1, VEGF-A and VEGF-C levels were measured before and during GH treatment in 6 patients with NS and 18 age-matched short subjects (Turner, idiopathic short stature and small for gestational age). MAIN OUTCOME MEASURES: Changes in plasma VEGF and IGF-1 levels. RESULTS: Baseline IGF-1 SDS levels were slightly lower in NS patients compared with controls; IGF-1 response to GH therapy was markedly lower in NS patients compared with controls (p = 0.017). Mean baseline VEGF-A levels were similar in NS patients and controls whilst mean baseline VEGF-C levels were significantly lower in the NS group as compared with controls (p = 0.022). Plasma VEGF-A and VEGF-C levels did not significantly change during GH treatment in the study cohort. No correlation was found between VEGF-C levels and levels of IGF-1, VEGF-A and auxological parameters, either before or during GH administration. CONCLUSION: Children with NS have a distinct growth factor profile including low basal VEGF-C and flattened IGF-1 response to GH. Further studies are needed to confirm our findings and to elucidate the interaction between VEGF-C levels and lymphedema.

Effects of a twelve-week randomized intervention of exercise and/or diet on weight loss and weight maintenance, and other metabolic parameters in obese preadolescent children.

AIMS: To compare the short- and long-term effects of intervention programs on body weight and cardiometabolic risk factors. METHODS: 162 obese children (6-11 years) were randomly assigned to three 12-week interventions with a 9-month follow-up period: exercise (E): 90 min moderate exercise 3 days/week (n = 52); diet (D): balanced hypocaloric diet, weekly meetings with dietician (n = 55), and diet + exercise (D+E) (n = 55). Changes in anthropometric variables, cardiometabolic profile and psychological outcome were assessed. RESULTS: At 12 weeks BMI-SDS, cardiometabolic profiles, and psychological score improved in all groups. The decrease in BMI-SDS was greater in D and D+E compared with E (p < 0.001), without a significant difference between the first two groups. Waist circumference and LDL cholesterol decreased more in D+E compared with E (p = 0.026 and p = 0.038, respectively). The increase in adiponectin was greater in D and D+E compared with E (p = 0.004). Anthropometric and cardiometabolic variables regressed without significant differences between groups after 9 months. However, BMI-SDS, body fat percentage and LDL cholesterol were lower compared to baseline. CONCLUSIONS: Diet alone or combined with exercise are the most effective short-term interventions for weight loss and improved cardiometabolic profiles, without a difference between them. In the long term, obese children need the long-term support of maintenance approaches.

Kisspeptin serum levels in girls with central precocious puberty.

OBJECTIVE: Central precocious puberty (CPP) causes early epiphyseal maturation, and early initiation of treatment improves final height. Unfortunately, there is no one parameter that can distinguish CPP from premature thelarche (PT), which is self-limited and requires no therapy. In animal models, kisspeptin, the ligand for the G-protein coupled receptor GPR54, was found to induce precocious activation of the gonadotrophic axis. Data on kisspeptin levels in girls with precocious puberty or in healthy prepubertal girls are lacking. We measured blood kisspeptin levels in girls with CPP and evaluated its potential as a clinical marker for CPP. Design This was a case-control study. PATIENTS: Thirty-one girls clinically diagnosed with CPP and 14 prepubertal age-matched healthy controls. Measurements Kisspeptin blood levels. RESULTS: Kisspeptin levels were significantly higher in the girls with CPP than in the controls: 14.62 +/- 10.2 pmol/l vs. 8.35 +/- 2.98 pmol/l, P < 0.05. Within the CPP group, there were no significant differences between the girls with a peak LH >5.0 IU/l and those with a peak LH

Nutrition-induced catch-up growth at the growth plate.

The effect of 40% food restriction (FR) and replenishment on the growth hormone (GH) and insulin-like growth factor-I (IGF-I) axis in the epiphyseal growth plate (EGP) was examined in a mouse model. Changes in RNA and protein levels were evaluated with real time PCR and immunohistochemistry, respectively, and serum levels of IGF-I and leptin were measured with radioimmunoassay. Dramatic changes in weight, tibial length and EGP height were observed following 10 days of 40% FR. The protein levels of IGF-I receptor (IGF-IR) and GH receptor (GHR), which were reduced during FR, increased during catch-up growth without an apparent change in the level of their RNA. The levels of type II and X collagens were unchanged. Serum IGF-I and leptin levels were reduced during FR and increased during catch-up growth. Following 40% FR, there was a significant decrease in the level of GHR and IGF-IR in the EGP which may explain the reduced effect of GH treatment in malnourished animals and children.

Leptin stimulates parathyroid hormone related peptide expression in the endochondral growth plate.

We have previously shown that growth plate chondrocytes expressed the long form of leptin receptor, and that within the growth plate, leptin stimulated cell proliferation and differentiation and epiphyseal growth in a balanced manner. These three cell processes are known to be regulated by the interactions of parathyroid hormone-related peptide (PTHrP) and Indian hedgehog (Ihh) protein. The aim of the present study was to examine the effect of leptin on the PTHrP/Ihh feedback loop. The effect of leptin was investigated in vivo in pair-fed experiments in ICR mice, and ex vivo in mandibular condyle explants incubated with leptin. Immunohistochemistry and in situ hybridization showed that in the pair-fed in vivo system as well as in the organ culture, leptin increased the level of PTHrP and reduced that of Ihh. Leptin may affect chondrocyte proliferation and differentiation by activating the PTHrP/Ihh growth-restraining feedback loop in the postnatal growth plate.

Leptin, insulin, insulin-like growth factors and their binding proteins in cord serum: insight into fetal growth and discordancy.

OBJECTIVE: Singleton infants with intrauterine growth restriction have an adaptive hormonal profile characterized by decreased levels of IGF-1, IGF-2, IGFBP-3 and insulin and elevated levels of IGFBP-1 and IGFBP-2. This study examined the association between cord serum levels of six growth factors and anthropometric features at birth in twins in order to determine the intrauterine growth factor interactions and to characterize the specific hormonal profile of small discordant twins. DESIGN: Prevalent case-control study. PATIENTS: Twenty pairs of discordant twins (5 monozygotic, 15 dizygotic) and 20 pairs of concordant twins (6 monozygotic, 14 dizygotic) matched for gestational age. MEASUREMENTS: Cord blood levels of IGF-1, IGF-2, IGFBP-1, IGFBP-3, insulin, leptin and anthropometric measurements at birth. Intra- and inter-pair differences and correlation coefficients were calculated, and the data were fitted to multivariate regression models. RESULTS: In both discordant and concordant groups, the smaller twins had a significantly lower level of IGF-1 (P < 0.03) and significantly higher level of IGFBP-1 (P < 0.02) than their larger cotwins. IGFBP-1 was inversely correlated with IGF-1 (P < 0.05). Insulin levels were significantly higher in the smaller discordant than the smaller concordant twins (P < 0.001) and in the larger discordant than the larger concordant twins (P < 0.004). Among the monozygotic twins, the leptin level was significantly higher in the larger discordant than the larger concordant twins (P < 0.025). Percentage birth weight discordancy was statistically correlated with twin-pair differences in IGF-1 and IGFBP-1. CONCLUSIONS: Of the six factors studied, IGF-1 appears to be the main indicator of intrauterine growth. Twin discordancy may involve compensatory rather than adaptive mechanisms or a multihormone relative resistance syndrome.

Effect of sex hormone administration on circulating ghrelin levels in peripubertal children.

BACKGROUND: Ghrelin levels gradually decrease throughout childhood and with advancing pubertal stage. The change during puberty is more pronounced in boys than girls. OBJECTIVE: The objective of the study was to investigate whether the pubertal drop in ghrelin secretion is modified by the increase in sex hormones. PATIENTS AND METHODS: Ghrelin levels were measured in 34 short peripubertal children (17 boys and 17 girls) aged 8-12.5 yr before and after sex hormone priming for GH stimulation testing. RESULTS: In boys, priming with testosterone increased testosterone to pubertal levels (23.7 +/- 7.1 nmol/liter), which in turn induced a marked decrease in ghrelin (from 1615.8 +/- 418.6 to 1390.0 +/- 352.0 pg/ml) and leptin (from 8.0 +/- 4.5 to 5.8 +/- 3.2 ng/ml) and an increase in IGF-I (from 162.7 +/- 52.8 to 291.1 +/- 101.6 ng/ml) (P < 0.001 for all parameters). In girls, priming with estrogen led to a supraphysiological increase in estradiol levels (1313.8 +/- 438.0 pmol/liter), which had no effect on ghrelin, leptin, or IGF-I. There was no correlation between ghrelin levels and levels of sex hormones, leptin, or body mass index in either boys or girls. CONCLUSIONS: A pharmacological increase in sex hormones is associated with a marked decline in circulating levels of ghrelin in boys but not girls. Additional longitudinal studies through puberty are needed to elucidate the physiological interaction between sex hormones and ghrelin.

Leptin reverses the inhibitory effect of caloric restriction on longitudinal growth.

Caloric imbalance, particularly in critical periods of growth and development, is often the underlying cause of growth abnormalities. Serum levels of leptin are elevated in obesity and are low in malnutrition and malabsorption. The aim of the present study was to determine whether leptin integrates energy levels and growth in vivo, as shown previously in our ex vivo experiments, even in the presence of caloric restriction. In the first part of the study, mice were divided into three groups. Two groups were fed ad libitum and received leptin or vehicle only, and the third group was pair-fed with the group injected with leptin to dissociate leptin's effect on growth from its effect on food consumption. Mice given leptin had a significantly greater tibial length than untreated pair-fed animals and a similar tibial length as control mice fed ad libitum despite their lower weight. In addition, leptin significantly increased the overall size of the epiphyseal growth plate by 11%. On immunohistochemistry and in situ hybridization studies, leptin stimulated both the proliferation and differentiation of tibial growth plate chondrocytes without affecting the overall organization of the plate. There was also a marked increase in the expression and level of IGF-IR. In the second part of the study, two groups of mice were fed only 60% of their normal chow; one was injected with leptin, and the other was injected with vehicle alone. Caloric deprivation by itself reduced serum levels of IGF-I by 70% and the length of the tibia by 5%. Leptin treatment corrected the fasting-induced growth deficiency, but further reduced the level of serum IGF-I. These results indicate that leptin stimulates growth even in the presence of caloric restriction independently of peripheral IGF-I.

Protective effect of insulin-like-growth-factor-1 against dopamine-induced neurotoxicity in human and rodent neuronal cultures: possible implications for Parkinson's disease.

Parkinson's disease (PD) is characterized by a progressive loss of 70-80% of dopaminergic (DA) neurons in the substantia nigra. High concentrations of DA were suggested to induce oxidative stress and selective neurodegeneration. We evaluated the effect of insulin-like-growth-factor-1 (IGF-1) on DA toxicity in neuronal cultures. IGF-1 (0.5 microg/ml) suppressed cell death induced by exposure to DA (0.3 mM) after 2 and 4 days, in a rat cerebellar culture. Similarly, IGF-1 (0.5 and 1.0 microg/ml) antagonized DA (0.125 and 0.250 mM) neurotoxicity in a human neuroblastoma cell line (SK-N-SH). Flowcytometric analysis of neuroblastoma cells treated with DA (0.5 mM) showed increased apoptosis, which was significantly reduced by IGF-1. The effect of IGF-1 was associated with increased Bcl-2 expression as indicated by flowcytometry and Western blot analysis. We suggest that IGF-1 possesses a neuroprotective effect against DA-induced toxicity, and may have a potential role in the treatment of PD.

Effect of dehydroepiandrosterone and its sulfate metabolite on neuronal cell viability in culture.

BACKGROUND: The neurosteroids dehydroepiandrosterone (DHEA) and its sulfated metabolite (DHEAS) have been reported to possess neuroprotective as well as anti-tumoral activity in vitro and in vivo. OBJECTIVES: To compare the effect of the two neurohormones on cell viability in primary whole-brain fetal mouse culture and isolated neuronal culture, as well as in a human neuroblastoma cell line (SK-N-SH). METHODS: Cell viability and cell proliferation were determined with the neutral red and 3H-thymidine uptake methods. Apoptosis in propidium iodide-stained neuroblastoma cells was determined using flow cytometry. RESULTS: DHEA (1 nM-10 microM) decreased the viability of selected primary neuronal cells (33-95% after 24 and 72 hours) but not of whole-brain cultured cells (neuron + glia). DHEAS did not significantly modify cell viability in either primary culture. In a human neuroblastoma cell line, DHEA (1 nM-1 microM) decreased 3H-thymidine uptake (30-60%) and cell viability (23-52%) after 24 hours. DHEAS did not significantly modify, or only slightly stimulated, cell viability and uptake of 3H-thymidine (132% of controls). The combination of DHEA and DHEAS neutralized the toxic effect of DHEA in both primary neuronal culture and neuroblastoma cell line. Flow cytometric analysis of DNA fragmentation in neuroblastoma cells treated with 100 nM DHEA/DHEAS for 24 hours showed an increase in apoptotic events (31.9% and 26.3%, respectively, vs. control 2.54%). CONCLUSIONS: Our results do not confirm a neuroprotective role for DHEA and suggest that DHEA and DHEAS have a differential role; DHEA possesses a neurotoxic (expressed only in isolated neurons) and anti-proliferative effect; DHEAS demonstrates only a slight neuroprotective effect.

Insulin, insulin-like growth factors-I and -II and insulin-like growth factor binding protein-3 in newborn serum: association with normal fetal head growth and head circumference.

The insulin-like growth factors (IGF) and their binding proteins (IGFBP) have been implicated in the regulation of fetal weight and length. The aim of our study was to determine the relationship between head circumference at birth and serum levels of IGF-I, IGF-II, IGFBP-3 and insulin in full-term appropriate-for-gestational age (AGA) infants. Serum samples were obtained from 77 singleton full-term neonates, 69 AGA and 8 small-for-gestational age (SGA). The AGA infants were divided into three groups by head circumference: Group 1: < or = 3rd percentile; Group 2: at 50th percentile; Group 3: > or = 97th percentile. Serum levels of IGF-I, IGF-II, IGFBP-3 and insulin were determined with commercial kits and immunometric methods. There were no statistically significant differences in mean serum levels of IGF-I, IGF-II and IGFBP-3 between the groups. A significantly higher mean serum insulin level was noted in the AGA infants with a head circumference > or = 97th percentile compared to those with a head circumference < or = 3rd percentile (4.6 +/- 0.3 vs 3.3 +/- 0.6 microU/ml; p = 0.04), and in AGA infants with a head circumference above the 50th percentile compared to those with a head circumference below the 50th percentile (4.4 +/- 0.4 vs 3.3 +/- 0.3 microU/ml; p = 0.01). AGA infants with a head circumference above or below the 50th percentile did not differ statistically in their mean IGF-II and IGFBP-3 serum level, while IGF-I differed statistically between the groups (18 +/- 2.7 vs 11.6 +/- 1.6 ng/ml, respectively; p = 0.045). Using univariate analysis, head circumference correlated positively with insulin (r = 0.29; p = 0.016) and with IGF-I (r = 0.26; p = 0.03). A stepwise multivariate linear regression analysis, however, did show statistically significant correlation of head circumference with birth weight (f = 36; p = 0.0001), and only marginally with birth length (f = 4.7; p = 0.06) and insulin (f = 3.4; p = 0.07). No correlations were found between head circumference and IGF-I, IGF-II or IGFBP-3. These data suggest that apart from genetic and nutritional factors, insulin may play a role in promoting intrauterine head growth, as reflected by head circumference at birth.

Evaluation of the neurotoxic activity of typical and atypical neuroleptics: relevance to iatrogenic extrapyramidal symptoms.

Typical neuroleptic therapy often results in extrapyramidal symptoms (EPS) and tardive dyskinesia (TD). Recent reports reveal neurotoxic activity in some neuroleptics. We hypothesized that neurotoxicity might be implicated in EPS. This study aims to evaluate the neurotoxic activity of typical and atypical neuroleptics and to determine the possible role of neurotoxicity in neuroleptic-induced EPS. Perphenazine, haloperidol, clozapine, sulpiride, and risperidone (10-100 microM) were administered, either alone or combined with dopamine, to primary mouse neuronal or intact brain culture and to a human neuroblastoma (NB) cell line (SK-N-SH). Cell viability (measured by neutral red and alamar blue), DNA fragmentation (flow cytometry-NB) were determined. Neuroblastoma: perphenazine, clozapine, and haloperidol (100 microM) decreased viability by 87, 43, and 34% respectively. Sulpiride and risperidone were not toxic. At 10 microM, toxicity decreased markedly. Dopamine (125 microM) potentiated the perphenazine-induced toxicity. Flow cytometry of NB cells treated with perphenazine (2.5-40 microM) showed an increase (perphenazine 20 microM, 40 microM, 48 h) in fragmented DNA (74.7% and 95.0% vs. 8.7% in controls). Lower concentrations increased the G1 phase and decreased S phase in the cell cycle. In primary neurons, perphenazine, haloperidol, and clozapine, but not risperidone and sulpiride, induced a significant neurotoxic effect, which, in intact brain culture, was absent (haloperidol and clozapine) or lowered (perphenazine). Dopamine (0.5 mM) did not modify the effect of the drugs in the primary cultures. Neuroleptics possess differential neurotoxic activity with higher sensitivity of neoplasm tissue (NB compared to primary cultures). The order of toxicity was perphenazine > haloperidol = clozapine:sulpiride and risperidone were not toxic. Neurotoxicity is independent of dopamine and is associated with cell cycle arrest and apoptosis. With the exception of clozapine, neurotoxicity seems relevant to neuroleptic-induced EPS and TD.

Basal plasma dehydroepiandrosterone sulfate level: a possible predictor for response to electroconvulsive therapy in depressed psychotic inpatients.

BACKGROUND: Dehydroepiandrosterone (DHEA) and its sulfate derivative DHEAS are neuroactive steroids. In the brain, they interact with gamma-aminobutyric acid (GABA(A)) receptors, which are involved in the regulation of anxiety and mood. The relevance of circulatory neurosteroids to psychiatric disorders and biological treatment is unknown. METHODS: Basal plasma levels of cortisol, DHEA, and DHEAS and the DHEAS-DHEA ratio were determined in 17 psychiatric inpatients before and after six electroconvulsive (ECT) therapy sessions, and all changes were statistically analyzed. For baseline values, 25 healthy individuals served as control subjects. Severity of depression and psychosis in the patients was measured with the Hamilton Depression Rating Scale (HDRS) and the Brief Psychiatric Rating Scale, respectively. RESULTS: Both basal and post-ECT levels of cortisol, DHEA, and DHEAS were significantly higher in the patients than in the control subjects. DHEAS levels in responding patients were higher at completion of treatment than at baseline. Patients defined as ECT nonresponders (change in HDRS < 30% from before treatments) exhibited elevated basal DHEAS levels compared with ECT responders. CONCLUSIONS: Markedly elevated basal DHEAS levels (mean + 2 SD of control value) are associated with resistance to ECT and may serve as a potential predictive marker of nonresponsiveness to ECT in depressed patients.

Elevated circulatory level of GABA(A)--antagonistic neurosteroids in patients with combat-related post-traumatic stress disorder.

BACKGROUND: Post-traumatic stress disorder (PTSD) is a multisystem neurobiological disorder with chronic alterations in various neurochemical systems. Levels of the GABA(A)--antagonistic neurosteroids plasma dehydroepiandrosterone (DHEA) and its sulphate derivate, dehydroepiandrosterone sulphate (DHEAS) may be relevant to depressive and anxiety disorders, including PTSD. METHODS: We assessed the circulatory levels of morning plasma DHEA and DHEAS in 21 male outpatients with untreated chronic combat-related PTSD (CR-PTSD), and 18 healthy control male subjects. RESULTS: Compared with the control subjects, the PTSD patients showed significantly higher plasma DHEA and DHEAS levels. CONCLUSIONS: Chronic CR-PTSD may be associated with increased circulatory level of neuroactive steroids with inhibitory activity at the GABA(A) receptors. Neurosteroid-induced decreased GABAergic tone may be relevant to the symptomatology and pathophysiology of chronic PTSD, as well as to the frequent co-morbidity of PTSD with depression and anxiety disorders.

Montelukast, a leukotriene receptor antagonist, reduces the concentration of leukotrienes in the respiratory tract of children with persistent asthma.

BACKGROUND: Leukotrienes are bronchoactive mediators secreted by inflammatory cells in the respiratory mucosa on exposure to asthma triggers. OBJECTIVE: We investigated the effect of montelukast, a leukotriene receptor antagonist, on the release of leukotrienes in the respiratory mucosa of children with persistent asthma. METHOD: Twenty-three children aged 6 to 11 years with moderately severe asthma were treated in a cross-over design starting, after a 2-week run in period, with either montelukast (n = 12) or cromolyn (n = 11) for 4 weeks with a 2-week washout period between treatments. Twelve of them were then treated with either montelukast or beclomethasone for 6 months. The use of beta(2)-agonists was recorded on a diary card. The concentration of leukotriene C(4) (LTC(4)) was measured by HPLC in nasal washes obtained before and at the end of each treatment period. Eosinophilic cationic protein (ECP) was measured in the nasal washes by RIA. RESULTS: The LTC(4) concentration significantly decreased in the children treated for the first 4 weeks with montelukast, from 5.03 +/- 1.17 to 1.42 +/- 0.33 ng/mL (P <.005), and a nonsignificant increase was noted in children treated with cromolyn, from 3.37 +/- 1.11 to 5.88 +/- 2.17 ng/mL (P =.17). ECP concentration also decreased in the children receiving montelukast (P =.12). The concentration of LTC(4) remained low after 3 and 6 months of treatment with montelukast (0.8 +/- 0.7 and 1.0 +/- 0.3 microg/mL) and was lower than with beclomethasone. Children treated with montelukast required significantly fewer beta(2)-agonists (P <.04), CONCLUSION: Montelukast reduces the concentration of leukotrienes in the respiratory tract of children with persistent asthma parallel to reduction in ECP and clinical improvement. This effect was not observed when the same children were treated with cromolyn.

Insulin-like-growth-factor-I (IGF-I) antagonizes apoptosis induced by serum deficiency and doxorubicin in neuronal cell culture.

We evaluated the effect of insulin-like growth factor (IGF)-I on neuronal cell viability and apoptosis induced by exposure to serum-free (SF) medium and to doxorubicin. In primary neuronal culture, IGF-I (0.5-2.0 microg/ml) slightly increased basal cell viability; SF medium tended to decrease viability (20-27%), and addition of IGF-I significantly antagonized this decrease (P< 0.05). In neuroblastoma (NB) SK-N-SH cell culture, IGF-I significantly increased viability (0.05-1.25 microg/ml) (P< 0.005); SF medium decreased it by 75%, and this decrease was prevented by IGF-I (0.5-1. 0 microg/ml) (P< 0.005). Flow cytometry studies showed an increased apoptosis on exposure to SF medium (88.8 vs 10.2%), which was suppressed to 38.3% by addition of IGF-I. Growth hormone (1-10 microU/ml) did not modify basal cell viability in either culture, and SF-induced cell death in NB cells. Doxorubicin (1-100 microM) caused neurotoxicity in primary and NB cultures (66-39% and 39-10% of controls, respectively), and increased apoptosis in NB cells (73. 8 vs 20.1%). IGF-I antagonized these neurotoxic/apoptotic effects (P< 0.05). This study suggests that IGF-I possesses a potent neuroprotective activity which may be involved in the resistance to doxorubicin.