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Osteocalcin (OCN), a marker of osteoblast activity, has been implicated in the regulation of energy metabolism by the skeleton and thus may affect body fat measures. We examined the relationships of OCN to body fat measures and whether they vary according to markers of energy and vitamin D metabolism. Data was obtained from 58 obese adolescents aged 13–17.9 years (38 females, 8 black or African-American). We calculated total fat mass (FM) [dual X-ray absorptiometry (DXA)] and visceral adipose tissue (VAT) [computerized axial tomography (CT)]. Blood tests included leptin, OCN, 25-hydroxyvitamin D [25(OH)D], parathyroid hormone (PTH), thyroid function tests, and triglycerides. Markers of glucose metabolism were obtained from fasting and OGTT samples. Adolescents with 25(OH)D

Background: Data on the relation between vitamin D status and body fat indexes in adolescence are lacking. Objective: The objective was to identify factors associated with vitamin D status and deficiency in obese adolescents to further evaluate the relation of body fat indexes to vitamin D status and deficiency. Design: Data from 58 obese adolescents were obtained. Visceral adipose tissue (VAT) was measured by computed tomography. Dual-energy X-ray absorptiometry was used to measure total bone mineral content, bone mineral density, body fat mass (FM), and lean mass. Relative measures of body fat were calculated. Blood tests included measurements of 25-hydroxyvitamin D [25(OH)D], parathyroid hormone (PTH), osteocalcin, type I collagen C-telopeptide, hormones, and metabolic factors. Vitamin D deficiency was defined as 25(OH)D 65 ng/mL. Results: The mean (±SD) age of the adolescents was 14.9 ± 1.4 y; 38 (66%) were female, and 8 (14%) were black. The mean (±SD) body mass index (in kg/m2) was 36 ± 5, FM was 40.0 ± 5.5%, and VAT was 12.4 ± 4.3%. Seventeen of the adolescents were vitamin D deficient, but none had elevated PTH concentrations. Bone mineral content and bone mineral density were within 2 SDs of national standards. In a multivariate analysis, 25(OH)D decreased by 0.46 ± 0.22 ng/mL per 1% increment in FM (β ± SE, P = 0.05), whereas PTH decreased by 0.78 ± 0.29 pg/mL per 1% increment in VAT (P = 0.01). Conclusions: To the best of our knowledge, our results show for the first time that obese adolescents with 25(OH)D deficiency, but without elevated PTH concentrations, have a bone mass within the range of national standards (±2 SD). The findings provide initial evidence that the distribution of fat may be associated with vitamin D status, but this relation may be dependent on metabolic factors. This study was registered at www.clinicaltrials.gov as {"type":"clinical-trial","attrs":{"text":"NCT00209482","term_id":"NCT00209482"}}NCT00209482, NCT00120146.

Iron-overload associated endocrinopathy is the most frequently reported complication of chronic transfusion therapy in patients with thalassaemia (Thal). This study compared iron-overloaded subjects with Thal (n = 142; 54%M; age 25.8 +/- 8.1 years) and transfused sickle-cell disease (Tx-SCD; n = 199; 43%M, 24.9 +/- 13.2 years) to non-transfused SCD subjects (non-Tx-SCD; n = 64, 50%M, 25.3 +/- 11.3 years), to explore whether the underlying haemoglobinopathy influences the development of endocrinopathy. Subjects were recruited from 31 centres in the USA, Canada and the UK. Subjects with Thal had more evidence of diabetes (13% vs. 2%, P < 0.001), hypogonadism (40% vs. 4%, P < 0.001), hypothyroidism (10% vs. 2%, P =

We report a case of a girl with clinical features of Peters' Plus Syndrome (PPS) (association of anterior eye chamber defects; peculiar facies; cleft lip/palate; brachymelia; developmental delay; growth retardation) and documented growth hormone deficiency (height -3.5 SDS at chronological age 5 years 8 months; low growth factors; bone age delay; growth velocity 4.4 cm/year (

Prader-Willi Syndrome (PWS), first described in 1956, is a unique genetic condition with a prevalence of 1 in 10,000 - 25,000. Features include severe lifelong hypotonia, insatiable appetite, short stature, obsessive-compulsive behaviour, morbid obesity, hypogonadism, kyphosis, scoliosis and osteoporosis. Studies beginning in the 1970s demonstrated that PWS is associated with a deficiency in growth hormone. Growth hormone treatment in children with PWS improves linear growth and more importantly leads to an increased muscle mass, bone mineral density and physical performance. In mid-2000, growth hormone became the first pharmaceutical approved in the US and Europe for the treatment of childhood PWS. It is now considered an essential part of comprehensive care for this condition. Ongoing studies address issues of growth hormone dosage, long-term efficacy, effects on neonatal and childhood growth and development and effects in adults with PWS.

Introduction: The liver is the main source of serum insulin-like growth factors (IGFs) and IGF-binding proteins (IGFBPs) and the concentration of these proteins might reflect liver function. Methods: In a retrospective longitudinal study we examined serum levels of total and free IGF-I, IGF-II, IGFBP-1, IGFBP-2, IGFBP-3 and IGFBP-6 in 21 adult patients with end-stage liver disease before and after orthotopic liver transplantation (LTX) by sensitive and specific RIAs. In each patient, the mean value of at least three measurements before and after LTX was calculated. Results: Before LTX, serum levels of total and free IGF-I, IGF-II, IGFBP-3 were low and showed a rapid and significant increase in almost all patients after successful LTX (total IGF-I: 30 ± 7 vs. 256 ± 30 ng/ml, p < 0.001; free IGF-I: 1.3 ± 0.3 vs. 3.5 ± 0.6 ng/ml, p < 0.01; IGF-II: 177 ± 28 vs. 618 ± 30 ng/ml, p < 0.001; IGFBP-3: 1,230 ± 136 vs. 3,665 ± 264 ng/ml, p < 0.001). In contrast, IGFBP-1 was found to be high immediately before LTX, and declined to normal levels after LTX (210 ± 40 vs. 90 ± 15 ng/ml, p < 0.01), while IGFBP-2 did not show any significant changes (1,154 ± 296 vs. 1,303 ± 192 ng/ ml). Positive correlations were found between IGF-I, IGF-II or IGFBP-3, and serum pseudocholinesterase (R = 0.50, 0.72 and 0.61 respectively, p < 0.001). Negative correlations were found between IGF-I, IGF-II or IGFBP-3, and prothrombin time (R = 0.50, 0.59 and 0.51 respectively, p < 0.001). Conclusion: Patients with severe liver disease show decreased levels of total and free IGF-I, IGF-II and IGFBP-3, and increased levels of IGFBP-1. These abnormalities are promptly normalized after successful LTX. Thus, serum levels of IGF-I, IGF-II and IGFBP-3 might be useful parameters for the assessment of liver function.

Prader-Willi syndrome (PWS) is a disabling condition characterized by hypotonia, hyperphagia, obesity, short stature, delayed or absent puberty, and mental retardation. The syndrome complex was first described in 1956 by Dr. Andrea Prader and colleagues [1]. In the 1980s, a characteristic genetic defect was identified involving deletion of paternal alleles at chromosome 15q11-13 [2-6]. This occurs by deletion of alleles on the paternal copy of chromosome 15q, an absent paternal chromosome 15q with maternal disomy, or, rarely, by mutations of the imprinting center of chromosome 15q. The estimated population prevalence of PWS is 1 in 15,000 live births.

Short stature, decreased muscle mass (hypotonia), increased body fat, decreased bone mineral density and other somatic abnormalities are major causes of morbidity and social limitation in individuals with Prader-Willi syndrome. Detailed studies indicate that two major endocrine pathologies may account for many of these somatic abnormalities. A true deficiency of the growth hormone (GH)-insulin-like growth factor axis is a principal cause of the short stature and is probably a major contributor to the decreased muscle mass and osteopenia. Hypogonadotropic hypogonadism is the probable primary cause of osteopenia and osteoporosis. No other endocrine abnormalities have been specifically identified in Prader-Willi syndrome, although there may be increased risks of premature adrenarche and type 2 diabetes mellitus, both secondary to obesity. GH replacement therapy is effective in normalizing linear growth and also has positive effects on muscle mass and function, and on bone mineralization. Judicious gonadal steroid replacement may be effective in treating the osteopenia and preventing osteoporosis. GH and gonadal steroid replacement therapy should be considered for all patients with Prader-Willi syndrome.

Children with chronic renal failure (CRF) are often growth recarded despite normal serum levels of GH and insulin-like growth factors (IGFs). Recent studies suggest that excess IGF-binding proteins (IGFBPs) in the 35-kDa fractions of CRF serum contribute to CRF growth failure. This report characterizes the relationship between IGFBP-3 and IGF peptides in the serum of growth-retarded CRF children. Size-exclusion chromatography at pH 7.4 found IGFBP-3 and IGFs almost exclusively in the 150-kDa fractions of normal serum, where their molar stoichiometry was approximately 1:1. However, similar chromatography of CRF serum found a molar excess of IGFBP-3 over total IGFs in the 150-kDa fractions and large amounts of IGFs in the 35-kDa fractions. In the 150-kDa fractions of CRF serum, IGFBP-3 was present in normal amounts, but a greater than normal amount was in the form of a 29-kDa IGFBP-3 fragment. Treatment of these CRF children with recombinant human GH increased the molar excess of IGFBP-3 over total IGFs in the 150-kDa fractions, the amount of IGFBP-3 and total IGFs in the 150-kDa fractions, and the amount of IGFs, but not IGFBPs, in the 35-kDa fractions. These data suggest that in untreated CRF children, proteolysis of IGFBP-3 in the 150-kDa fractions releases IGFs to the excess IGFBPs in the 35-kDa fractions, but insufficient IGF is released to overcome the growth-inhibiting effects of these excess IGFBPs. Treatment with recombinant human GH increases levels of IGFs and IGFBP-3 in the 150-kDa fractions, and subsequent IGFBP-3 proteolysis releases sufficient IGF to overcome the growth inhibitory effects of excess IGFBPs in the 35-kDa fractions of CRF serum.

The activity of the hypothalamic-GH-insulin-like growth factor (IGF) network declines with age. It has recently been shown that increased cardiovascular mortality occurs in adults with GH deficiency. As hypercholesterolemia is common in GH-deficient adults, and because there is experimental evidence that GH may play a role in regulating plasma cholesterol, we decided to investigate the activity of the GH-IGF axis in an elderly population by measuring serum IGF-I, IGF-II, and IGF-binding protein-3 (IGFBP-3) levels and to study their relationship with blood lipid levels. One hundred and thirty-two elderly subjects, 52 men and 80 women, were studied (age range, 60–91 yr). Men had significantly lower levels of IGFBP-3, high density lipoprotein cholesterol (HDL-C) and apoprotein A1 (ApoA1) compared to the women, whereas IGF-I and IGF-II were only slightly lower. Using linear regression analysis, we observed an inverse relationship of age with IGF-I (r = −0.35; P < 0.001), IGF-II (r = 0.40; P < 0.001), IGFBP-3 (r = 0.52; P < 0.001), body mass index, and lipid levels. Univariate regression analysis showed a strong and positive correlation of both IGF-I and IGFBP-3 with HDL-C and ApoA1. Partial correlation analysis, after adjustment for age and body mass index, showed that IGFBP-3 and IGF-II were still significantly and positively related to HDL-C and ApoA1. Furthermore, a strong association was documented among IGF-I, IGF-II, and IGFBP-3. These data demonstrate that even in an elderly population, further aging is accompanied by a progressive decline in circulating IGF-I, IGF-II, and IGFBP-3, suggesting a continuing diminution of the GH-IGF axis throughout aging. Moreover, the strong correlation between HDL-C and an index of GH secretion, such as IGFBP-3, suggests that GH might play an important role in lipid metabolism in healthy elderly subjects.

Previous studies suggest that growth retardation in children with chronic renal failure (CRF) results in part from inhibition of insulin-like growth factor (IGF) action by excess serum IGF-binding proteins (IGFBPs). Excess IGFBPs in CRF serum include IGFBP-1, -2, and -3 and a diffuse approximately 24- to 28-kDa IGFBP band identified by [125I]IGF ligand blot. The present studies characterized this diffuse approximately 24- to 28-kDa band. Initial studies identified this band as IGFBP-6, because it was immunoprecipitated by antiserum raised against a synthetic peptide of human IGFBP-6 (hIGFBP-6). Additional [125I]IGF ligand blots found that the immunoprecipitated band was 1) recognized by [125I]IGF-II but not [125I]IGF-1, 2) more abundant in CRF than in normal serum, and 3) more abundant in serum from dialyzed than nondialyzed prepubertal CRF children. Using the hIGFBP-6 antiserum in a specific and sensitive RIA, we found that serum IGFBP-6 levels were 4.7 +/- 1.7 nmol/L in 10 normal prepubertal children, 21.4 +/- 6.1 nmol/L in 44 nondialyzed prepubertal CRF children, 73.5 +/- 14.4 nmol/L in 7 dialyzed prepubertal CRF children, and 94.6 +/- 26.2 nmol/L in 14 dialyzed pubertal CRF children. IGFBP-6 levels were also elevated in 71 nondialyzed European children with CRF. In nondialyzed CRF children, serum IGFBP-6 levels 1) correlated inversely with the glomerular filtration rate, 2) did not correlate with height SD score, and 3) were not altered by 12 months of daily recombinant hGH treatment. In summary, a specific antiserum and RIA were used to demonstrate elevated levels of intact IGF-II-binding IGFBP-6 in serum of CRF children. We postulate that the excess IGFBP-6 may modulate the action of IGF-II on target tissues.

Children with chronic renal failure (CRF) have normal or high serum levels of GH, IGF-I, and IGF-II. Despite this, the serum of CRF patients has low IGF bioactivity, which may contribute to CRF growth failure. Recent studies suggest that excess IGF binding proteins (IGFBPs) in the approximately 35-kD fractions of CRF serum contribute to this low IGF bioactivity. This report characterizes a 29-kD form of IGFBP-3, IGFBP-3(29), which accumulates in the approximately 35-kD fractions of CRF serum and peritoneal dialysate. Deglycosylation and [125I]IGF ligand blot studies show that IGFBP-3(29) is a glycosylated IGFBP-3 fragment with low affinity for IGF peptides. Using an IGFBP-3 antibody column, IGFBP-3(29) was purified to homogeneity from the approximately 35-kD fractions of peritoneal dialysate from children with CRF. Compared with native IGFBP-3, pure IGFBP-3(29) has a 4-10-fold lower affinity for IGF-II and a 200-fold lower affinity for IGF-I. Consistent with the binding data, IGFBP-3(29) inhibited IGF-II-stimulated thymidine incorporation in chondrosarcoma cells, but was a less potent inhibitor than native IGFBP-3; also, native IGFBP-3 clearly inhibited IGF-I-stimulated thymidine incorporation in chondrosarcoma cells and potentiated IGF-I-stimulated aminoisobutyric acid uptake in bovine fibroblasts, but higher concentrations of IGFBP-3(29) had no effect on these IGF-I actions. Thus, the 29-kD IGFBP-3 form that accumulates in CRF serum and extravascular spaces is an IGFBP-3 fragment that may modulate IGF-II, but not IGF-I, effects on target tissues. Whether IGFBP-3(29) plays any role in the growth failure of children with CRF remains to be determined.

OBJECTIVE: In normal subjects, the major form of circulating IGF is the GH-dependent 150 kDa complex. The liver appears to be the main source of the three components of the 150 kDa complex and, in particular, hepatocytes synthesize the insulin-like growth factor (IGF) peptide and the acid-labile subunit (ALS), whereas Kupffer and sinusoidal endothelial cells produce IGF-binding protein-3 (IGFBG-3). We have studied the effects of the somatostatin analog octreotide, IGF-II des(1-3)IGF-I, transforming growth factor (TGF)-beta 1 and tri-iodothyronine (T3) on ALS secretion into the medium conditioned by rat hepatocytes in primary culture. METHODS: The regulation of ALS release was evaluated in the conditioned medium of adult rat hepatocytes exposed to increasing concentrations of test substances or to vehicle alone (control), after gel filtration in basic conditions, by immunoblot using an antiserum generated against the N-terminal 34 amino acids of human ALS. RESULTS: The results demonstrate that: 1) octreotide in vitro produces a dose-dependent inhibition of both basal and GH-stimulated ALS secretion into the hepatocyte conditioned medium; 2) the release of ALS by adult rat hepatocytes is not affected by the presence during the incubation of des(1-3)IGF-I or IGF-II; 3) an inhibitory effect, although only with very high doses, can be observed after treatment with TGF-beta 1; and 4) a small but significant increase of ALS released into the medium can be seen when hepatocytes are treated with T3. CONCLUSIONS: Evaluation of the effect of substances known to affect the production of IGF peptides, the IGFBPs, or both, on adult rat hepatocytes in primary culture revealed no powerful stimulator, but instead a potent inhibitor of ALS release/synthesis. Our data suggest that the effect of somatostatin on the 150 kDa complex is mediated not only by the reduction in GH concentration, but also by a direct inhibition of ALS release or synthesis.

The in vivo physiological relationships among GH, the insulin-like growth factors (IGFs), and the IGF-binding proteins (IGFBPs) are not completely defined, and single random measurements of these serum proteins do not completely reveal their dynamic relationships. We report the kinetic responses of the IGFs and IGF-binding proteins to exogenous GH in 23 subjects with untreated GH deficiency [5 women and 18 men; age, 15.0 +/- 6.2 yr (+/- s.d.), height z-score = -4.4 +/- 2.2 (+/- s.d.); body mass index = 19.3 +/- 2.4 kg/m2]. After an overnight fast, subjects were given a sc dose of recombinant human GH (2.85 i.u./m2), and blood was sampled from an indwelling peripheral venous catheter 0, 1, 2, 3, 4, 5, 6, 8, 10, 12, 14, and 24 h after the injection. Subjects were then treated with recombinant human GH (2.85 i.u./m2.day); fasting samples were obtained at 3 months (n = 22), and timed sampling was repeated at 6 months (n = 21). Fasting levels of IGF-I, free IGF-I, IGF-II, IGFBP-3, and insulin increased significantly within 3 months of GH treatment, whereas IGFBP-1, IGFBP-2, and IGFBP-6 showed no change. In the timed sampling studies at 0 and 6 months, GH levels peaked 3 h after treatment; the degree of rise and the rate of decline were both greater at 6 months. IGF-I levels increased beginning at 4 h, continuing throughout the 24-h period at month 0, whereas a plateau was observed after 6-8 h during the 6-month study. Free IGF-I paralleled total IGF-I except during fasting, when it varied inversely with IGFBP-1. IGFBP-3 and IGF-II both showed late (> 20 h) responses to a dose of GH, whereas IGFBP-2 and IGFBP-6 showed minimal changes. IGFBP-1 varied inversely with insulin, which, in turn, varied with meal intake. Comparative studies in 2 subjects with GH receptor deficiency showed no response to exogenous GH. However, both IGFBP-1 and IGFBP-2 were several-fold elevated, and IGFBP-1 varied inversely with the low insulin levels. Our data are the first to examine multiple elements of the serum IGF system in response to GH in both GH-deficient and replete states. The relationships of the different response patterns provide insight into the physiology of this system and may guide future studies.

Modulation of growth factors by growth hormone in children with chronic renal failure. Anthropometric measurements and circulating growth factors were studied serially in 44 prepubertal children with growth failure and chronic renal failure (GFR = 10 to 40 ml/min/1.73m2) who were randomized to receive either recombinant human growth hormone (rhGH; N = 30) or no treatment (N = 14). RhGH was given as Nutropin, 0.05 mg/kg/day, and the studies were carried out at baseline and after 3 and 12 months. At baseline, serum insulin-like growth factor binding protein (IGFBP)-1 and -2 levels were, while IGFBP-3 levels were not, higher than those of children with normal renal function. In addition, height SDS at baseline correlated inversely with serum IGFBP-2 levels (r = -0.461, P = 0.0016), but did not correlate significantly with any other factor. After 12 months of study, the 30 children receiving rhGH showed: (i) greater increase in height (9.1 ± 2.8 vs. 5.5 ± 1.9 cm, P < 0.0001); (ii) increases in serum levels of IGF-I, IGF-II, free IGF-I, IGFBP-3 and acid labile subunit (ALS); (iii) a greater decrease in serum IGFBP-1 levels; and (iv) no significant difference in serum IGFBP-2 levels, when compared to the 14 control patients. The change in height SDS after 12 months of rhGH (+ 0.8) in the 30 treated children correlated significantly and positively with serum ALS, IGFBP-3, total IGF, IGF-I, IGF-II and free IGF-I levels measured during treatment. These observations suggest that, in children with growth failure associated with chronic renal failure: (i) IGFBP-2, and not IGFBP-3, is likely to be a growth inhibitor; (ii) rhGH stimulates catch-up growth in part by increasing serum levels of IGF peptides; and (iii) linear growth is influenced by the balance between growth stimulating IGFs and growth inhibitory IGFBPs.

Weight loss is a common, persistent characteristic of long term human immunodeficiency virus (HIV-1) infection; its full etiology remains unknown. Because treatment with GH has induced nitrogen retention in various catabolic conditions, we designed this study to determine whether a moderate dose of insulin-like growth factor I (IGF-I) combined with a low GH dose could impede the catabolic response seen in HIV-1 infection. A double blind, placebo-controlled study design was used. Subjects in the GH/IGF-I treatment group (n = 44) and control group (n = 22) continued to receive their routine stable antiretroviral therapy. No patient had a recent history of opportunistic infection, malignancy, or Kaposi's sarcoma and had dietary intakes of at least 25 Cal/kg weight.day at study entry. During the 12-week study period, dietary instruction was given, and subjects were encouraged to maintain an intake of 35 Cal/kg and 1 g protein/kg. All subjects had a body mass index of 19.8 kg/m2 or less at the time of study entry or a weight loss of 10% or more of their premorbid weight and a body mass index below 26.1 kg/m2. The treatment group received 0.34 mg (0.68 mg/day) GH, twice daily, and 5.0 mg (10 mg/day) IGF-I, twice daily. Changes in body composition of total body potassium (TBK), total body nitrogen (TBN), fat-free mass (FFM), and body fat (Fat) were examined at 6 and 12 weeks during the treatment period. TBK, TBN, FFM, and Fat for the treatment and placebo groups were, on the average, below normal at study entry. At 6 weeks, the GH/IGF-I group showed a significant increase in FFM (P0.0001), a minimal increase in TBK (P0.05), and a substantial decrease in Fat (P0.01) compared with baseline values. The loss of body fat continued to be significant (P0.01) in the GH/IGF-I group treatment at 12 weeks, whereas the increase in FFM was minimal (P0.05). No significant changes in the mean body composition occurred at 6 or 12 weeks in the placebo group. By 12 weeks, neither TBK (body cell mass) nor TBN (total protein mass) had significantly increased relative to the values at baseline, although the FFM remained elevated. Thus, the combined GH and IGF-I doses used in this study in adult males with HIV-associated weight loss were ineffective in producing a sustained anabolic response and, in fact, resulted primarily in a significant loss of body fat.

Children with chronic renal failure (CRF) often fail to attain an adult height consistent with their genetic potential. The growth hormone (GH)/insulin-like growth factor (IGF)/growth plate chondrocyte axis has been intensively studied in these children to determine the basis for this growth failure. Evidence suggests that hepatic GH resistance results in deficient expression of IGF-I. However, serum IGF-I levels are usually normal and it is IGF-I action on target tissues which is inhibited, possibly by the presence of excess high-affinity IGF binding proteins (IGFBPs) in CRF serum. In this paper we evaluate the roles of IGFBP-1, -2, and -3 as growth inhibitors in CRF children. The data support a role for each of these IGFBPs as growth inhibitors. Currently, IGFBP-1 meets most criteria expected of a growth inhibitor, but IGFBP-2 and -3 will likely also meet these criteria and may well be important contributors to the growth failure of CRF. Ultimately, many or all of the six IGFBPs may be found to contribute to the excess high-affinity IGF binding sites which are a hallmark of CRF serum and are possible contributors to the growth failure of CRF children.

We report the development of a unique radioimmunoassay (RIA) for rat insulin-like growth factor-I (rIGF-I) which does not recognize human IGF-I (hIGF-I). The rIGF RIA uses a specific anti-rIGF-I antiserum with rIGF-I standards and radioligand. Rat sera were extracted by either an abbreviated acid/ethanol procedure or by acid-chromatography with virtually identical results. Assay sensitivity is congruent to 0.5 ng/tube and inter- and intra-assay coefficients of variation were 3.4-7.6% and 4.8-9.2%, respectively. Comparisons of the rIGF-I RIA with a typical heterologous RIA shows that the latter underestimates rIGF-I levels congruent to of 3-fold. Sera from rats treated with hIGF-I were also studied using the rIGF-I RIA and a hIGF-I- specific immunoradiometric assay (IRMA), and results indicate poor correlation between the actual total IGF-I levels (rIGF-I RIA + hIGF-IRMA) and the heterologous RIA estimates. The availability of both rIGF-I and hIGF-I specific immunoassays facilitates more precise studies of hIGF-I in the rat model.

Insulin-like growth factors I and II (IGF-I and II) are present in serum primarily within a ternary complex consisting of IGF, IGF-binding protein-3 (IGF-3) and acid-labile subunit (ALS). Relatively little is known about ALS as compared to the other components of the complex. We report immunoblot studies of ALS using a new rabbit antiserum to human ALS1-34. The antiserum shows high specificity for ALS, labelling only the intact 82-88 kDa doublet in whole serum. Treatment with endoglycosidase-F leads to only a partial deglycosylation of ALS in whole serum, while purified ALS is reduced to M(r) approximately 58 kDa. Acidification of both whole serum and purified ALS leads to a complete loss of ALS ability to bind to cross-linked IGFBP-3:[125I]IGF-II tracer; however, immunoblot studies show no change in the apparent M(r) of the major ALS band. Immunoblot studies of human serum shows that intact ALS is decreased in growth-hormone (GH) deficiency, increases with GH treatment, is elevated in GH excess and is unchanged with IGF-I treatment. These data provide new information regarding the characteristics of ALS and demonstrate the research utility of a highly-specific antiserum for this protein.

Insulin-like growth factor-binding protein-1 (IGFBP-1) is one of several related proteins that bind and modulate the actions of IGFs. The liver is the primary source of IGFBP-1, and insulin is a major regulator of hepatic IGFBP-1 production. We report five sets of investigations that further define the characteristics of hepatic IGFBP-1 response to insulin. In normal subjects, a continuous high-dose insulin infusion caused a rapid decrease in plasma IGFBP-1 concentrations, with a rate of 0.24 ± 0.04 μg/L·min−1 and a t 1 2 of 89 ± 4 minutes. Conversely, a 3-hour somatostatin (SRIF) infusion caused a 4.5-fold increase in plasma IGFBP-1 levels. SRIF plus low-dose insulin infusion (to inhibit break-through insulin secretion) resulted in a plateau in IGFBP-1 concentrations at 5 to 8 hours, with a t 1 2 to achieve steady state of 60 to 75 minutes. Under similar conditions, a stepped increase in plasma glucose level from 5 to 9 mmol/L had no effect on the rate of IGFBP-1 increase in plasma, indicating that an acute increase in glucose concentration within a physiologic range has no independent inhibitory effect on IGFBP-1 production in the presence of a nonsuppressive insulin level. Using SRIF plus sequential graded insulin infusions, the threshold peripheral (=portal) plasma insulin concentration for IGFBP-1 suppression was between 65 and 172 pmol/L. Subjects with insulin-dependent diabetes mellitus (IDDM) showed a similar dose-response pattern, suggesting that insulin regulation of IGFBP-1 may be normal in IDDM. The rapid and sensitive response of IGFBP-1 to insulin is consistent with its postulated role to modulate IGF bioaction in relation to substrate availability. Moreover, measurement of plasma IGFBP-1 concentrations may serve as a useful marker of hepatic insulin sensitivity.

Insulin-like growth factor binding protein-1 (IGFBP-1) is a liver-derived protein that modulates the mitogenic actions of the insulin-like growth factors (IGFs). IGFBP-1 production is potently inhibited by insulin both in vivo and in HepG2 human hepatoma cells. To further define the pathways of IGFBP-1 regulation, we studied the effects of modulators of protein kinase-C (PKC) on HepG2 cell IGFBP-1 production. Phorbol 12-myristate 13-acetate (PMA) stimulated IGFBP-1 production in a time- and dose-dependent manner, with maximal stimulation occurring at 10-100 nmol/L. The degree of stimulation was dependent on cell density, ranging from about 2-fold in confluent to more than 10-fold in sparse cultures. Preincubation with PMA abolished the inhibitory effect of insulin, while preincubation with insulin did not inhibit PMA stimulation. The transient PKC activator diC8 had no effect, while studies with the PKC inhibitors sphinganine and H-7 were limited by solvent vehicle cytotoxicity. Staurosporine (STS), a potent PKC inhibitor, stimulated IGFBP-1 production 2- to 4-fold and augmented the stimulatory effect of PMA. Concanavalin-A, an inhibitor of PMA-stimulated PKC translocation and down-regulation, inhibited the effects of PMA and STS. Our findings indicate that PKC is involved in the regulation of hepatic IGFBP-1 production. The effects of PMA, which causes rapid activation, followed by membrane translocation and down-regulation of PKC, are similar to those of STS and are countered by Concanavalin-A. These data suggest that PKC activity may mediate tonic inhibition of IGFBP-1 production, while PKC downregulation stimulates the production of this regulatory protein.

Insulin is the principal regulator of hepatic insulin-like growth factor binding protein-1 (IGFBP-1) production, mediating the rapid decrease in plasma IGFBP-1 in response to nutritional intake. In this study, we defined IGFBP-1 regulation by insulin in upper and lower body obesity, conditions associated with insulin resistance and chronic hyperinsulinemia. Overnight postabsorptive IGFBP-1 levels in obese and nonobese women showed an inverse, nonlinear relationship with plasma insulin concentrations. Maximum suppression of IGFBP-1 was seen at 70-90 pmol/L plasma insulin. Both groups of obese women had mean fasting plasma insulin concentrations above this threshold level and, consequently, markedly suppressed IGFBP-1 levels. To assess the dynamics of insulin regulated IGFBP-1, 10 obese and 8 nonobese women were studied during sequential saline infusion (0-90 min), hyperinsulinemia (insulin infusion; 90-210 min) and hypoinsulinemia (somatostatin + GH infusion; 210-330 min). Insulin infusion rapidly decreased plasma IGFBP-1 levels in nonobese subjects (60% decrease in 2 h), but had little or no further suppressive effect in obese subjects. Complete insulin withdrawal resulted in a significant rise in plasma IGFBP-1 concentrations in all subjects, but the response was blunted in obese compared to nonobese groups. In contrast to plasma IGFBP-1, IGF-I concentrations did not vary during hyper- and hypoinsulinemic infusion periods and were not significantly different between groups. Basal GH levels were significantly higher in nonobese when compared to obese women, but did not change with infusions. In conclusion, low IGFBP-1 levels in obesity are related to elevated insulin levels which are, in turn, related to body fat distribution and insulin resistance. The chronically depressed levels of IGFBP-1 may promote IGF bioactivity as well as its feedback regulation of GH secretion, thus contributing to the metabolic and mitogenic consequences of obesity. In addition, our findings imply that hepatic insulin sensitivity in terms of IGFBP-1 production is preserved despite peripheral insulin resistance in obesity.

We examined the relationships between 4 glycated protein assays and home blood glucose monitoring (HBGM) in 26 children with poorly-controlled insulin-dependent diabetes mellitus (IDDM) during a period of improved management. At 2 week intervals for 6 visits (12 weeks in total), HBGM records were collected and a blood sample was obtained for measurement of glycated proteins and glucose. Assays included glycated hemoglobin (GHb) and glycated serum proteins (GP) by boronate affinity chromatography, hemoglobin A 1 C by PolyCAT A high performance liquid chromatography (HAC) and fructosamine (FA). All 4 glycated protein levels declined significantly over the 12 week period. Significant correlations between the glycated proteins and HBGM were observed over 2 week intervals. None of the 4 assays were affected by the glucose level in the sample. Changes in mean HBGM readings over 2 week intervals were correlated with both FA and GP with wide prediction intervals. Over cumulative 2 week intervals, which may more accurately reflect longitudinal trends, all 4 glycated proteins were correlated with mean HBGM readings. At each cumulative interval, GHb and GP showed the largest variation with MBG, while FA showed the least variation with MBG. Our data indicate that of the 4 assays tested, FA has limited clinical value as compared to other glycated protein assays, whereas assays based on boronate affinity chromatography (GHb and GP) provide the most useful clinical indicators of short-term changes in glycemic control. The clinical utility of a new HPLC method for determination of glycated hemoglobins is also demonstrated.

Insulin rapidly lowers serum insulin-like growth factor-binding protein-1 (IGFBP-1) levels in vivo. In studies reported here, HEP G2 cells were used as a model system to investigate how insulin achieves this effect. When HEP G2 cells were incubated with 100 nM insulin for 6, 14, or 24 h, IGFBP-1 protein levels in conditioned medium fell to approximately 50% of control values. This apparently was due to a fall in the rate of IGFBP-1 protein synthesis, since HEP G2 cells incorporated 46% less [35S]methionine into IGFBP-1 during a 4-h incubation with 100 nM insulin. IGFBP-1 mRNA levels were similarly affected by 100 nM insulin, falling to 45% of control values after 2 h, and to 9% of control values after 4 h of incubation with this hormone. The fall in IGFBP-1 mRNA level is consistent with data from nuclear transcription assays. HEP G2 nuclei isolated from cells that were incubated with 100 nM insulin for 2 h synthesized only approximately 1/3 the number of IGFBP-1 transcripts as did control nuclei. Further evidence that insulin decreases IGFBP-1 gene transcription comes from transient transfections using chimeric IGFBP-1 promoter-chloramphenicol acetyltransferase reporter gene constructs. IGFBP-1 promoter activity fell to approximately 50% of control values when HEP G2 cells transfected with a construct containing the first 1205 base pairs of the IGFBP-1 promoter were incubated with 100 nM insulin for 6, 14, or 24 h. Insulin lowered both IGFBP-1 protein levels and promoter activity in a dose-dependent manner. A half-maximal effect was found at approximately 1 nM insulin and a maximal effect was found at approximately 10 nM insulin in each instance. Transfections with constructs containing smaller IGFBP-1 promoter fragments showed that the region spanning from 103 to 529 base pairs 5' to the IGFBP-1 mRNA cap site was necessary to demonstrate the inhibitory effect of insulin. These studies indicate that insulin lowers IGFBP-1 protein levels, at least in part, by rapidly decreasing the rate of IGFBP-1 gene transcription, and suggest that this insulin-mediated fall in transcription is conferred through a specific region of the IGFBP-1 promoter.

GH replacement in Prader-Willi syndrome (PWS) children has well-defined benefits and risks and is used extensively worldwide. Its use in PWS adults has been limited by documentation of benefits and risks, as determined by larger multisite studies.Our objective was to evaluate the effectiveness and safety of GH in GH-deficient genotype-positive PWS adults.We conducted a 12-month open-label multicenter trial with 6-month dose-optimization and 6-month stable treatment periods.The study was conducted at outpatient treatment facilities at four U.S. academic medical centers.Lean and obese PWS adults with diverse cognitive skills, behavioral traits, and living arrangements were recruited from clinical populations.Human recombinant GH (Genotropin) was initiated at 0.2 mg/d with monthly 0.2-mg increments to a maximum 1.0 mg/d, as tolerated.Lean body mass and percent fat were measured by dual-energy x-ray absorptiometry.Lean body mass increased from 42.65 +/- 2.25 (se) to 45.47 +/- 2.31 kg (Por = 0.0001), and percent fat decreased from 42.84 +/- 1.12 to 39.95 +/- 1.34% (P = 0.025) at a median final dose of 0.6 mg/d in 30 study subjects who completed 6-12 months of GH. Mean fasting glucose of 85.3 +/- 3.4 mg/dl, hemoglobin A1c of 5.5 +/- 0.2%, fasting insulin of 5.3 +/- 0.6 microU/ml, area under the curve for insulin of 60.4 +/- 7.5 microU/ml, and homeostasis model assessment of insulin resistance of 1.1 +/- 0.2 were normal at baseline in 38 study initiators, including five diabetics, and remained in normal range. Total T(3) increased 26.7% from 127.0 +/- 7.8 to 150.5 +/- 7.8 ng/dl (P = 0.021) with normalization in all subjects, including six (20%) with baseline T(3) values at least 2 sd below the mean. Mildly progressive ankle edema was the most serious treatment-emergent adverse event (five patients).This multicenter study demonstrates that GH improves body composition, normalizes T(3), and is well tolerated without glucose impairment in PWS genotype adults.

Insulinlike growth factor binding protein 1 (IGFBP-1) has been shown to modulate the metabolic and mitogenic actions of the growth hormone (GH)- dependent peptide insulinlike growth factor I. Previous studies showed that levels of IGFBP-1 are regulated by insulin. The relative role of GH in the regulation of IGFBP-1 levels is less well defined and was examined in our study with a contiguous two-part protocol. Overnight (part A) and pre- and post-morning meal (part B) blood samples were obtained from eight healthy adults during a constant infusion of saline (SAL) or 4 μg · kg−1 · min−1 GH. Five of eight subjects were restudied with glucose (GLUC) infused during part B (SAL + GLUC) to match glucose and insulin to levels observed during GH infusion. During SAL infusion, IGFBP-1 levels measured by specific radioimmunoassay showed a marked immediate decline after the evening meal in part A, with a subsequent nocturnal rise of 2.4- to 17.3-fold. GH infusion resulted in a similar meal-induced fall in IGFBP-1 levels but led to a delayed nocturnal rise in IGFBP-1, which was associated with elevated postprandial insulin concentrations. During part B, changes in plasma IGFBP-1 levels showed a similar pattern, with a delayed postprandial increase observed during both GH and SAL + GLUC infusions. The half-life of IGFBP-1 disappearance was calculated at ∼2 h for all three infusion groups. Comparison of venous and arterialized blood samples showed no consistent pattern of difference, arguing against peripheral tissue clearance or compartmentalization as the mechanism for the rapid rise and fall in IGFBP-1 levels. Our studies, the first to measure GH, insulin, and glucose in concurrent sampling and under controlled physiological conditions, support previous investigations suggesting that insulin (not GH or glucose) is the primary regulator of plasma IGFBP-1 levels. Furthermore, we postulate that the observed fluctuations in plasma levels are caused by a direct insulin effect on hepatic IGFBP-1 production.

A human hepatoma cell line, HepG2, secretes a discrete insulin-like growth factor-binding protein (IGFBP-1) into serum-free medium, which is identical to the 25K mol wt BP in amniotic fluid and plasma. IGFBP-1 levels in vivo have been shown to be inversely correlated with circulating insulin concentrations. This study investigated the direct effects of insulin on IGFBP-1 production in vitro. Addition of insulin to HepG2 cultures induced a rapid dose-dependent decrease in IGFBP-1 synthesis and secretion independent of the glucose concentration in the medium. As assessed by ligand binding and specific RIA, levels of IGFBP-1 were 20-50% of control levels in 18-h conditioned medium from insulin-treated cells. Monoclonal antibody studies indicated that the suppressive effect of insulin on IGFBP-1 synthesis was mediated through specific interaction with the insulin receptor. Therefore, HepG2 cells respond to insulin by altering the synthesis and secretion of IGFBP-1 in a manner that mimics many of the changes in plasma IGFBP-1 levels observed in vivo and provide an in vitro model for studies of IGFBP-1 biosynthesis.

Isolated growth hormone deficiency (IGHD) represents conditions of GH deficiency that are not necessarily associated with other pituitary hormone deficiencies or with an organic lesion. Three sub-categories of IGHD have been clinically identified (IGHD types 1–3), and IGHD type 1 has been further separated into IGHD types 1a and b. However, this clinical sub-categorization of IGHD may need reconsideration due to the recent identification of molecular heterogeneity within each sub-type of IGHD. In a small number of children with IGHD, defects in the GH, GH-releasing hormone receptor (GHRH-R), and GH1 genes have been identified. In most cases, no cause for IGHD can be identified; however, the proportion of idiopathic IGHD cases may be decreasing due to identification of causative factors. The phenotype of IGHD is variable depending in part on the underlying genetic disorders in the affected individuals. Several studies have focused on the usefulness of MRI findings in patients with GHD but anatomic abnormalities of the pituitary gland are variable. We review current studies and the clinical, biochemical, and molecular features described for different groups of affected individuals with IGHD.

Pediatric pituitary diagnosis is complicated by the unique developmental characteristics, secretory patterns and regulation of each pituitary hormone system. Although dynamic test procedures have been described, validation of each procedure is limited and universally agreed diagnostic criteria are not available. Clinical acumen, experience and judgment continue to be primary elements of pituitary diagnosis.

Objective The mouse serves as an important model for insulin-like (IGF)-related research. However, lack of homologous mouse assays has prevented studies of the normal ontology of the murine IGF system. Therefore, we developed and validated immunoaassays for murine IGF-I, IGFBP-3 and ALS and studied levels of these analytes in mice. Methods Using commercially available reagents, we developed and validated specific enzyme-linked immunosorbent assays (ELISAs) for murine IGF-I, IGFBP-3, and ALS. Levels of these analytes were measured in sera from CD-1 mice, male and female, sampled at 1, 2, 4, 8, 20 and 32 weeks of age. In addition, sera from pregnant and postpartum CD-1 mice were also studied. Results Validation of specific ELISAs for murine IGF-I, IGFBP-3 and ALS are described; all 3 assays were highly sensitive, precise and accurate. As measured by our homologous ELISA, IGF-I levels (ng/mL, mean ± SD) in male mice were relatively low at 1 week (53 ± 8), rising sharply to peak at 8 weeks of age (636 ± 48), and gradually declining thereafter, reaching 395 ± 64 at 32 weeks. IGF-I levels in non-pregnant female mice peaked at 4 weeks of age (548 ± 77) declined at 8 weeks (417 ± 61), then recovered to plateau at 539 ± 78 and 535 ± 88 at 20 and 32 weeks, respectively. In male mice, trends in IGFBP-3 were similar to the patterns of IGF-I. However, in non-pregnant female mice, the IGFBP-3 levels declined relatively slowly after peaking at 4-weeks of age, unlike IGF-I levels during this period. ALS levels followed the same pattern as IGF-I in both sexes. IGF-I to IGFBP-3 molar ratios (percent) were similar between sexes, rising continuously with age: ∼30% at 1 week, 80% at 4 weeks, 135% at 32 weeks. IGF-I was reduced in 8 week old mice in mid-pregnancy (354 ± 75 vs 417 ± 61 in non-pregnant 8 week females), reaching a nadir in late-term (146 ± 40), and only partially recovering in the postpartum period (239 ± 23). IGFBP-3 was also lower in late-pregnancy (1245 ± 100 vs 1925 ± 439) and remained depressed postpartum. In contrast to IGF-I and IGFBP-3, ALS increased more than threefold in mid-pregnancy (12180 ± 1641 vs 3741 ± 910), followed by a 4-fold decrease in late-pregnancy (2964 ± 489), recovering postpartum (6104 ± 1178). Conclusions We report the first ontological studies of IGF-I, IGFBP-3 and ALS in mice using newly-characterized sensitive, homologous immunoassays. Our results indicate that mice have a generally similar pattern in IGF-related axis components, with low levels early in life, increasing to peak during sexual maturation and declining thereafter. Significant gender differences in non-pregnant levels and dramatic changes during pregnancy were also found. Knowledge of the normal developmental changes in the murine IGF system and accurate tools for investigations of this system are a necessary foundation for research in this field.

Pituitary gigantism, a condition of endogenous growth hormone (GH) hypersecretion prior to epiphyseal closure, is a rare condition. In the adult condition of GH excess, acromegaly, the occurrence of type 2 diabetes mellitus (T2DM) and diabetic ketoacidosis (DKA) have been reported, with resolution following normalization of GH levels. We report the case of a 16-year-old male with pituitary gigantism due to a large invasive suprasellar adenoma who presented with T2DM and DKA. Despite surgical de-bulking, radiotherapy and medical treatment with cabergoline and pegvisomant, GH and insulin-like growth factor-I (IGF-I) levels remained elevated. However, the T2DM and recurrent DKA were successfully managed with metformin and low-dose glargine insulin, respectively. We review the pathophysiology of T2DM and DKA in growth hormone excess and available treatment options.

Insulin-like growth factor-binding protein-1 (IGFBP-1) is one of six soluble binding proteins that regulate the actions of the insulin-like growth factors (IGFs). Liver is the major source of IGFBP-1 in non-pregnant humans. In normal physiology, IGFBP-1 transcription is potently inhibited by insulin and serum levels are limited by a rapid clearance rate. Elevated levels of IGFBP-1 in liver disease have been attributed to insulin resistance; however, the relationships between these analytes have not been defined. We studied insulin, proinsulin and IGFBP-1 in normal subjects (NL, N=47, 43+/-12 yr), cirrhosis (CIR, N=29, 54+/-14 yr), hepatocellular carcinoma (HCC, N=42, 61+/-11 yr), and other liver tumors (TUM, N=8, 60+/-17 yr). All three analytes were significantly increased in liver disease (mean+/-SEM; p-values relative to normals): IGFBP-1 (NL 24+/-4 ng/ml; CIR 235+/-53, p

Cells are known to undergo apoptosis when cultured in high serum concentrations. However, the serum factors responsible for this induction of apoptosis have not been identified. The IGF-binding protein-3 (IGFBP-3), a negative growth regulator, is found at concentrations of 5 microgram/ml in serum. We have recently demonstrated that IGFBP-3 induces apoptosis in PC-3 cells, a prostate cancer cell line, at a concentration of 500 ng/ml. In this communication, we demonstrate the role of IGFBP-3 as one of the apoptosis-inducing agents in high serum concentrations. Treatment of PC-3 cells with increasing concentrations (40% to 90%) of intact human serum (HS) resulted in a dose-dependent decrease in cell growth. Valinomycin, an ionophore, was used as a positive control to measure the induction of apoptosis by serum treatment in PC-3 cells. Treatment with 90% serum showed significant suppression of growth (P

The possible occurrence of increased non-enzymatic glycosylation of serum insulin-like growth factor binding protein-3 (IGFBP-3) in vivo and the changes that would simultaneously occur in serum levels of IGFBP-3 and insulin-like growth factor-1 (IGF-I) were investigated. We measured levels of IGF-I and IGFBP-3 and the degree of glycation of total serum protein and IGFBP-3, in serum samples obtained from patients with poorly controlled non-insulin-dependent diabetes (type 2) and from age-matched non-diabetic controls. Type 2 diabetic patients had significantly higher glycated serum protein (GlyP) levels. GlyP significantly correlated with age in the control (r = 0.315, P, Facultad de Ciencias Médicas

IGFBPs play an important role in IGF biological actions by modulating IGF binding to its receptors. The major IGFBP in serum is IGFBP-3, which transports 70-90% of the circulating IGFs. In target cell systems, it sequesters IGFs and inhibits their hormonal actions, but may potentiate IGF activity or exert IGF-independent effects under specific conditions. IGFBP-3 can be modified by IGFBP-3 proteases, which degrade it into smaller fragments. IGFBP-3 fragments generated by proteolysis have reduced affinity for IGFs, thereby modifying IGF action. To study IGFBP-3 fragments in vivo and in vitro, we constructed six different IGFBP-3 fragments by use of a baculovirus expression system and generated 8 different monoclonal IGFBP-3 antibodies. Based on the known cleavage sites of IGFBP-3 for PSA, MMPs, and the predicted plasmin cleavage sites, we expressed a N-terminal IGFBP-3(1-97) fragment and a C-terminal IGFBP-3(98-264) fragment. By stepwise truncation from the C-terminal end, we created IGFBP-3(98-232), IGFBP-3(98-206), IGFBP-3(98-179), and IGFBP-3(98-159). A strong recognition of the C-terminus and the intermediate parts of IGFBP-3 by six antibodies was found. Four of these mAbs were able to recognize the intermediate fragment alone. Two mAbs were found to immunoreact only with the N-terminal IGFBP-3 fragment and two additional mAbs recognized the N- as well as the C-terminal parts and lacked immunoreactivity for the intermediate part of IGFBP-3. The 15 kDa IGFBP-3 fragment resulting from plasmin digestion was found to only react with N-terminal antibodies, while the 29 kDa fragment in pregnancy serum reacted with both N- and C-terminal antibodies. Thus, these mAbs will be useful tools to determine whether IGFBP-3 fragments found in vivo derive from either the N- or C-terminal domains of IGFBP-3.

3 alpha-androstanediol glucuronide (3 alpha diolG) is a marker of peripheral tissue androgen metabolism. There are no previous data regarding complete paediatric reference ranges for 3 alpha diolG. In order to obtain reference values for 3 alpha diolG we have measured serum levels of 3 alpha diolG in 283 healthy children and adolescents, 146 boys and 137 girls, age 1 month to 20 years and 28 adults. A non-extraction, solid phase radioimmunoassay employing a polyclonal antiserum that is specific for 3 alpha diolG was used to measure serum 3 alpha diolG levels (intra assay variation 5.1-10.1%, inter assay variation 2.7-9.0%). There was a strong sex and age dependence (r = 0.8; p < 0.0001) of 3 alpha diolG levels throughout childhood and adolescence with males showing significantly higher levels of the androgen than females (p < 0.05). 3 alpha diolG serum levels (nmol/l +/- SD) correlated significantly with pubertal stage (p < 0.01). Interestingly, in 35 children with CAH serum 3 alpha diolG levels correlated well with clinical and metabolic status, i.e. 17OHP serum levels. In summary, we have established percentile curves for 3 alpha diolG levels in healthy children and adolescents. We hypothesize that on the basis of our reference values the single measurement of serum 3 alpha diolG could serve as a means to determine androgen status in children with disorders of puberty and sexual development.

Serum insulin-like growth factors (IGFs), which circulate bound to specific IGF binding proteins (IGFBPs), must exit the intravascular space before acting on target tissues. Little is known about the nature of IGF/IGFBPs in extravascular fluids of patients with chronic renal failure (CRF). Peritoneal dialysate (PD) was studied since, after a short incubation, PD contains proteins which have entered an extravascular space; thus, IGF/IGFBP forms in PD are more likely than serum forms to interact with target tissues. IGF-I and IGF-II, and IGFBPs 1 – 4, were readily identified by specific immunoassays and/or 125iodine-IGF ligand blotting of simultaneously obtained PD and serum samples from seven CRF children; IGFBP-3 was a major IGFBP in PD as in serum. Where quantitated, IGF and IGFBP levels in PD were approximately 10% of serum concentrations. After separation of PD and serum by size-exclusion chromatography, serum had more IGFBP-3 in 150-kilodalton (kDa) than 35-kDa fractions, while PD had far less IGFBP-3 in 150-kDa than 35-kDa fractions. Immunoblot studies revealed a major 29-kDa IGFBP-3 fragment, in addition to intact 41- and 38-kDa IGFBP-3 forms, in PD and CRF serum; the 29-kDa form predominated in the 35-kDa PD fractions. These data suggest that the 29-kDa fragment is the IGFBP-3 form most likely to modulate IGF effects on target tissues of CRF individuals.

Insulin-like growth factors (IGFs) and their binding proteins (IGFBPs) have important regulatory functions in ovarian follicular development. Although most studies have investigated the IGF system in ovarian cells in vitro, investigation of the IGF system in the peripheral circulation and in follicles of varying sizes throughout the menstrual cycle in large numbers of subjects has been lacking. In the current study we performed daily IGF-I, IGF-II, IGFBP-1, and IGFBP-3 measurements in 9 healthy regularly cycling volunteers throughout the menstrual cycle. In addition, we investigated IGF-I, IGF-II, IGFBP-1, and IGFBP-3 levels in 13 samples of androgen-dominant follicular fluid [FFa androstenedione to estradiol (AD:E2) ratio, > 4] and 19 samples of estrogen-dominant follicular fluid (FFe; AD:E2 ratio, 4) obtained from 21 regularly cycling subjects and in 18 samples of fluid from luteinizing follicles obtained from patients undergoing in vitro fertilization (IVF) treatment (FFivf). IGF-I, IGF-II, IGFBP-1, and IGFBP-3 were measured using two-site immunoradiometric assays. No significant day to day differences were observed in IGF-I, IGF-II, IGFBP-1, and IGFBP-3 levels across the menstrual cycle. Median IGF-II levels in FFe (630 ng/mL; range, 212-1000) were significantly higher compared to those in FFa (474 ng/mL; range, 272-603; P = 0.002). Median IGFBP-3 levels in FFe (2955 ng/mL; range, 388-3448) were also significantly higher than those in FFa (2352 ng/mL; range, 756-2604; P = 0.003). Median IGF-I (192 ng/mL; range, 29-256) and IGFBP-1 (12 ng/mL; range, 2-281) levels in FFe were not significantly different from those in FFa [149 (range, 22-232) and 21 (range, 5-32) ng/mL, respectively). In contrast, significantly lower IGFBP-1 levels were found in FFe compared to FFivf (79 ng/mL; range, 57-234; P = 0.002), whereas there was no significant difference between FFe and FFivfe IGF-I, IGF-II, or IGFBP-3 levels, respectively. IGF-II levels were correlated with follicle diameter (r = 0.52; P = 0.002), cycle day (r = 0.47; P = 0.0065), E2 levels (r = 0.53; P = 0.003), AD:E2 ratio (r = -0.58; P = 0.001), and P concentrations (r = 0.60; P = 0.001) in all follicles, whereas no such correlations were found with IGF-I. In conclusion, as circulating levels of IGF-I, IGF-II, IGFBP-1, and IGFBP-3 are not menstrual cycle dependent, it is unlikely that these growth factors and these binding proteins play an endocrine role in cyclic ovarian follicle development, although both cycle-dependent delivery to the ovary and modification of their actions locally within the ovary cannot be excluded. With regard to FF1 the findings that IGF-II levels in FF1 are elevated compared to those in FFa and correlate with follicular functional status support a role for IGF-II during development of the dominant follicle. In addition, as IGFBP-3 in estrogen-dominant follicles mirrors the rise of IGF-II, this IGFBP may be a primary regulator of IGF-II action within the estrogen-dominant follicle. Finally, the finding of elevated levels of IGFBP-1 in luteinizing (IVF) follicles suggests an important role for this peptide in corpus luteum regulation.

The regulation of insulin-like growth factor binding protein-1 (IGFBP-1) by its ligands, IGF-I and IGF-II, was studied in continuous cultures of HepG2 human hepatoma cells. Both IGF-I and IGF-II in concentrations as low as 1–10 nmol/l caused significant suppression of IGFBP-1 protein levels. This suppression was accompanied by decreased IGFBP-1 mRNA levels occurring within 2–4 h of exposure to IGF-I or IGF-II, and by a significant decrease in IGFBP-1 promoter activity. IGF-I and IGF-II were equipotent in suppressing basal levels of IGFBP-1 protein, mRNA and promoter activity. IGF-I, IGF-II, and IGF-analogs with low IGFBP-1 affinity, (des 1–3)IGF-I and long R3IGF-I, all potently suppressed the previously characterized increase in IGFBP-1 protein levels and promoter activity induced by cAMP and theophylline. In contrast, [Leu-27]IGF-II, which interacts with the type II but not type I IGF receptor, had no effect on IGFBP-1 protein levels or promoter activity. Our data indicate that IGFBP-1 production is inhibited by its ligands, IGF-I and IGF-II, and that this effect is probably mediated at the transcriptional level. The effects of IGF-I and IGF-II apparently occur as a result of binding to the type I IGF receptor, and are similar to the previously characterized suppressive effects of insulin on IGFBP-1 transcription mediated through the insulin receptor. When considered with previous data regarding expression of IGFBP-1 and the type I IGF receptor, our results suggest that IGF regulation of IGFBP-1 may play an as yet undefined role in fetal development and postnatal hepatic regeneration.

Prader-Willi Syndrome (PWS) is characterized by infantile failure to thrive, progressive childhood obesity, hypotonia, hypogonadism, mental retardation and phenotypic abnormalities including short stature. Abnormalities of chromosome 15 can now be identified in virtually all cases1. The pathogenesis of PWS is not known, although the phenotype suggests that there may be an abnormality in substrate utilization leading to an absolute deficit of lean mass and increased fat mass. Hypothalamic and pituitary abnormalities have also been postulated2,3. However, studies of body composition, nutrient utilization, and hormone status in PWS are limited. We report initial results from a detailed investigation of body composition and growth hormone (GH) treatment in PWS.

Clinical testing of growth hormone (GH) sufficiency is a controversial area in endocrinology. Due to the episodic nature of endogenous GH secretion, diagnosis of GH deficiency has been defined as a failure to achieve normal GH levels in response to at least two stimuli. This testing is associated with significant patient morbidity and cost. We analyzed our experience over a 4-y period to determine whether clinical or biochemical variables could be used to predict the results of a specific GH testing procedure. Of 180 cases analyzed (67% male, mean age 8.89 +/- 4.39 y, range neonate-16 y), eight cases had incomplete GH testing results. Of the remaining 172, 19 were GH deficient (GH level less than 7 ng/mL). Younger age, higher body mass index and a greater degree of bone age delay were characteristic of the GH-deficient population; however, none of these variables alone was of diagnostic utility. Serum IGF-I level was below the normal range for 81% of the GH deficient and 47% of the GH-sufficient children; and was the only single variable that provided a reasonable between-group distinction. Discriminant analysis resulted in development of a new variable, based on IGF-I z scores, chronologic age, degree of bone age delay, and body mass index, which would have allowed exclusion of GH deficiency without provocative testing for 58% of the GH sufficient population, whereas permitting the diagnosis of GH deficiency for all GH-deficient subjects. Our data are dependent on the IGF-I assay method and the clinical definition for GH deficiency; therefore, the calculated predictive values are not applicable to all clinical populations.(ABSTRACT TRUNCATED AT 250 WORDS)

From a physiologic point of view, the somatic tasks that are unique to the first two decades of life are linear growth, primarily due to bone lengthening, and sexual maturation. The key control centre for both of these processes is the pituitary, the smallest, most complex and most fragile of all discrete multifunctional endocrine glands. Arguably, our understanding of the phylogeny of the pituitary and, in particular, the evolutionary pathways leading to its situation as a unitary ductless secretory organ in chordates, has progressed minimally since a seminal review in 1924 [1]. In addition, as briefly reviewed in Chapter 2, longstanding controversies in clinical diagnosis and treatment of childhood pituitary disorders remain unresolved. However, over the past few years, advances in molecular genetics have contributed to a burgeoning understanding of both normal and abnormal pituitary function. This special issue of Pituitary is intended to provide an overview of the pituitary as it pertains to pediatric medicine. The issue begins with a short historical introduction authored by Dr. Solomon Kaplan, a pioneer in pediatric endocrine research. Clinical controversies and the interesting condition of pituitary gigantism are then briefly discussed, before moving into a focus on recent progress in molecular biology, with sections authored by leading investigators in their respective fields. We wish to express our gratitude to the authors for their contributions of time, effort and knowledge, both in the preparation of this issue and, more importantly, in furthering our understanding of pituitary physiology. We also thank Dr. Shlomo Melmed for the invitation to prepare this issue.

Sir .—The nonmedical use of anabolic steroids among high school students is an issue that requires better definition. Terney and McLain 1 present data that show that 4.4% of the students in a large Chicago, Ill, high school admit to using anabolic steroids, an incidence that is quite concerning. However, the interpretation of the data is compromised by questions regarding the study design. In my experience, the term anabolic steroid is relatively unfamiliar to many, if not most, school-aged children. None of the survey questions in this study verified that the subjects understand this term, and no mention was made of presurvey education. An unknown percentage of the children will be familiar with the term steroid as used for treatment of asthma, allergies, and other medical conditions and might answer in the affirmative without knowing the meaning of anabolic . Another unknown percentage of the children may have had anabolic

Plasma samples from 68 growth hormone (GH)-deflclent children (provocative serum GH level

Changes in the normal hormonal regulation of linear growth which lead to growth failure in children with chronic renal failure (CRF) are not well understood. Previous studies indicate that serum levels of growth hormone and IGF-I and II are normal or elevated in this population; and that serum levels of poorly defined inhibitors of IGF action are increased. Using a recently developed RIA for the 25-kD IGF-binding protein (IGF-BP25), we report significant elevations of this protein in children with CRF when compared to age- and sex-matched controls. IGF-BP25 levels correlate positively with IGF-binding activity in both populations, indicating that the RIA reflects levels of bioactive protein. Although the variation of serum IGF-BP25 with chronologic age and IGF levels are preserved in CRF, the quantitative interrelationships are disrupted. Levels of the 53-kD IGF-binding protein, an IGFbinding protein derived from the high mol wt IGF complex, were also found to be elevated in the CRF population and, unlike in the control population, did not vary with age or IGF levels. IGF-BP25 has been shown to inhibit IGF mitogenic action in vitro. Our finding of elevated levels of IGF-BP25 in children with CRF suggests that this protein may play a role in the growth retardation of pediatric chronic renal failure. The significance of the elevated IGFBP53 levels in CRF remains uncertain.

Corticosterone methyl oxidase deficiency type II (CMO type II deficiency) is an autosomal recessive disorder characterized by a defect in the terminal step of aldosterone biosynthesis. Previous reports emphasized the diagnostic utility of elevated urinary ratios of 18-hydroxytetrahydro-compound A to tetrahydro-aldosterone, which are primary metabolites of 18-hydroxycorticosterone (18-OHB) and aldosterone, respectively. Limited data suggest that plasma ratios of 18-OHB to aldosterone are also abnormal in affected individuals. We report serum steroid profiles in two siblings with CMO type II deficiency. Serum levels of aldosterone precursors were elevated in both patients before treatment. In particular, the serum ratios of 18-OHB to aldosterone were greatly elevated and declined to normal levels during mineralocorticoid replacement. The possibility of heterozygote detection using this ratio is suggested. We also confirm previous reports of a detrimental effect on linear growth rate after cessation of mineralocorticoid therapy despite maintenance of normal serum electrolytes. This effect is associated with biochemical evidence of chronic salt depletion. Based on our observations, we recommend that serum 18-OHB to aldosterone ratios be routinely measured for the diagnosis and management of patients with CMO type II deficiency.

Previous investigations have demonstrated specific receptors and associated mitogenic actions for insulin and insulinlike growth factors I and II (IGF-I and II) in postnatal bovine aortic smooth muscle. Using fetal tissue we have observed different patterns of binding and action for these peptides. Smooth muscle cells isolated from near-term fetal bovine aortae were studied in early passage. Specific receptors for both IGF-I and IGF-II were identified. Specific binding averaged 5.7%/2.5 X 10(5) cells for IGF-I, and 16.2% for IGF-II, and 0.3% for insulin. High affinity Kd for both IGF receptors were nanomolar. IGF-II was fivefold less potent than IGF-I in displacing IGF-I binding. IGF-I showed no affinity for the IGF-II receptor. Insulin, at physiologic concentrations, was incapable of displacing either IGF-I or IGF-II binding. Cellular incorporation of [methyl-3H]thymidine was stimulated at the lowest dose of IGF-I tested, 0.5 ng/ml. IGF-II showed no effect up to 100 ng/ml, after which a sharp increase in incorporation was noted. Insulin had a similar effect only at concentrations greater than 0.5 micrograms/ml, with a maximal response noted at 5 to 10 micrograms/ml. Our results indicate that fetal bovine aortic smooth muscle cells have an abundance of IGF receptors but lack specific insulin receptors. In addition, IGF-II binding levels are three times higher than for IGF-I. These results are consistent with observations in other species, in which a predominance of IGF over insulin receptors has been demonstrated in fetal tissue, and provide further evidence for a role for the IGFs in embryonic cellular metabolism.

Nucleotide sequencing of the two known cDNAs encoding human insulin-like growth factor I (IGF-I) predicts two different prohormone forms of IGF-I. The predicted prohormone amino acid sequences (E peptide regions) extend the carboxy-terminus of IGF-I by either an additional 35 (IGF-IA) or 77 (IGF-IB) amino acids. We developed an antiserum directed against a synthetic peptide which is unique to the E peptide region of IGF-IA prohormone. In a RIA using this antiserum, synthetic E peptide immunoreactivity was found in the serum of patients with chronic renal failure. The protein recognized by this antiserum has a mol wt of about 13,000 by neutral gel filtration and about 19,000 by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoblotting. These data suggest that the E peptide region of the IGF-IA prohormone is expressed and circulates as part of the prohormone.