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<0.0001; HCC 505+/-105, p<0.0001; TUM 118+/-36, p<0.0001), insulin (NL 72+/-4 pM; CIR 261+/-62, p<0.0002; HCC 180+/-25, p<0.0001; TUM 189+/-58, p<0.0001), proinsulin (NL 6.5+/-0.7 pM; CIR 36.8+/-7.7, p<0.0001; HCC 26.2+/-3.8, p<0.0001; TUM 32.1+/-9.7, p<0.0001). The ratio of proinsulin to insulin was also significantly elevated in liver disease. A typical curvilinear inverse relationship of insulin and IGFBP-1 was observed, but was shifted several fold higher for the liver disease groups. Our results demonstrate that insulin and proinsulin are elevated in liver disease. However, these elevations are paradoxically accompanied by elevated IGFBP-1 levels, indicating disruption of normal regulatory mechanisms. IGFBP-1 is postulated to play a dynamic role in metabolic substrate utilization via regulation of free IGF. Therefore, inappropriate elevation of IGFBP-1 could play an important role in the metabolic disturbances associated with liver disease.

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 is a defining feature of PWS [1, 7]. Children with PWS typically have a gradually declining linear growth velocity beginning in early childhood [8-10]. Virtually all adults with PWS are significantly below their midparental height, with an average adult height approximately 2 SD below the general population average. This degree of short stature creates a significant barrier to social adaptation and performance.

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.

Polycystic ovary syndrome (PCOS) is the most common cause of anovulation in women.Previous studies suggest that the pathogenesis of PCOS may involve interrelated abnormalities of the insulinlike growth factor (IGF) and ovarian steroidogenesis systems.We investigated this hypothesis in fasting serum samples from 140 women with PCOS (age, 27.4 Ϯ 0.4 yr; body mass index, 26.3 Ϯ 0.5 kg/m 2 ; mean Ϯ SEM).IGF-related parameters were also studied in a group of normoovulatory women (n ϭ 26; age, 26 Ϯ 4 yr; body mass index, 23.6 Ϯ 4.3 kg/m 2 ).For the PCOS group, the mean testosterone (T) level was 2.5 Ϯ 0.1 nmol/L, and it was significantly correlated with LH (r ϭ 0.41; P Ͻ 10 Ϫ6 ), estrone (r ϭ 0.33; P ϭ 0.016), estradiol (r ϭ 0.18; P ϭ 0.04), and androstenedione (AD; P Ͻ 10 Ϫ6 ), but not with dehydroepiandrosterone sulfate (P ϭ 0.71), a marker of adrenal steroidogenesis.T and AD were also related to total ovarian follicle number and ovarian size, as previously found with normoovulatory women (1).There were no differences between the PCOS subjects and the normoovulatory group for total IGF-I, IGF-II, or IGF-binding protein-3 (IGFBP-3).However, IGFBP-1 levels were significantly decreased in the PCOS group (1.0 Ϯ 0.2 vs. 7.3 Ϯ 1.1 ng/mL; P Ͻ 0.001) and were inversely correlated with serum insulin levels (r ϭ Ϫ0.50; P Ͻ 10 Ϫ8 ).Serum levels of free IGF-I (fIGF-I) were elevated (5.9 Ϯ 0.3

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.

Children with chronic renal failure (CRF) are often growth retarded 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.

BACKGROUND:Previous studies indicate that resting energy expenditure is elevated in children with sickle cell anemia, possibly caused in part by hemolysis and increased erythropoietic activity. The purpose of the present investigation was to determine whether erythrocyte transfusion normalizes resting energy expenditure in sickle cell anemia.METHODS:Five adolescents with sickle cell anemia (12-16 years old; 4 boys, 1 girl) were studied before and 1 week after erythrocyte transfusion before elective surgery or at the initial transfusion for growth failure. Resting energy expenditure was measured by indirect calorimetry, and laboratory measures were determined by routine, validated methods. Data comparisons were by nonparametric analysis.RESULTS:After erythrocyte transfusion, total hemoglobin levels increased (difference (D) = 15 g/l; p < 0.05), whereas hemoglobin S (D = -0.36; p < 0.05) and reticulocyte count (D = -0.12; p < 0.05) decreased. Mean pretransfusion resting energy expenditure was elevated to 124% above predicted levels (p < 0.05) and increased further to 134% above prediction (p < 0.05 vs. pretransfusion levels). Plasma triiodothyronine (T3) levels increased (D = 0.17 nmol/l; p < 0.05), reverse T3 (rT3) levels tended to decline (D = -0.04 nmol/l; p = 0.14), and rT3/T3 decreased (D = -0.03; p < 0.05). Plasma insulin-like growth factor-I (IGF-I) levels were low-normal before transfusion and did not change, despite the change in resting energy expenditure.CONCLUSIONS:The results confirm that resting energy expenditure is elevated in patients with sickle cell anemia. However, resting energy expenditure further increased after transfusion, despite decreased erythropoietic activity. A posttransfusion decrease in rT3/T3 may contribute to the increased resting energy expenditure. That there was no change in IGF-I implies that the growth hormone-IGF system is not involved in posttransfusion regulation of resting energy expenditure. Therefore, our data are not consistent with the hypothesis that increased resting energy expenditure in sickle cell anemia is directly related to erythropoietic activity. The mechanisms by which resting energy expenditure increases after transfusion in sickle cell anemia require additional investigation.

Injury to the rotator cuff tendons is a common cause of pain and disability in the upper extremity. The supraspinatus tendon is frequently torn, either as a direct result of trauma alone or through trauma following attritional changes. When surgical repair is performed, patients all too often regain only limited shoulder motion. One probable reason for lingering stiffness is that active shoulder rehabilitation is delayed for weeks following surgery for fear of disrupting the repair. Recent research aimed at hastening the healing of muscle, ligaments, and tendons through the application of biological agents called growth factors focuses on boosting natural repair processes, thereby facilitating accelerated rehabilitation and return to function. Less long-term stiffness can be expected when earlier active motion is undertaken, with the end result being both an earlier and a more functional return to unrestricted activity. In this article, we propose a new approach to accelerating functional recovery following rotator cuff repair, using exogenous application of biological agents to boost tendon healing.

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.

The insulin-like growth factors (IGF-I and IGF-II) and their binding proteins (IGFBPs) have been implicated in regulating fetal growth and development. The aim of this study was to determine whether fetal IGFs correlate with auxologic data at birth and/or gestational age. Venous cord blood was obtained from 138 healthy newborns immediately after birth and clinical data were recorded using a standardized data sheet. For the determination of IGF-I and IGF-II, IGFBP-blocked radioimmunoassays were used, A coated-tube immunoradiometric assay was applied for the measurement of free IGF-I. IGFBP-1, -2, and -3 were measured using specific radioimmunoassays. IGF-I levels were 61 +/- 21 ng ml(-1), median 61 ng ml(-1), range 19-114 ng ml(-1); IGF-II levels were 466 +/- 80 ng ml(-1), median 457 ng ml(-1), range 311-701 ng ml(-1); free IGF-I levels were 2.4 +/- 1.8 ng ml(-1), median 1.8 ng ml(-1), range 0.4-7.8 ng ml(-1). The concentration of IGFBP-1 was 144 +/- 110 ng ml(-1), median 113 ng ml(-1), range 20-626 ng ml(-1); that of IGFBP-2 was 1165 +/- 455 ng ml(-1), median 1119 ng ml(-1), range 440-3466 ng ml(-1). IGFBP-3 levels were 1272 +/- 280 ng ml(-1), median 1272 ng ml(-1), range 600-1966 ng ml(-1). IGF-I levels correlated significantly with IGFBP-3 levels (r = 0.71), birthweight (r = 0.48) and birth length (r = 0.37). There were significant inverse correlations between IGF-I and both IGFBP-1 (r = -0.45) and IGFBP-2 (r = -0.62). Although free IGF-I levels correlated (r = 0.71) with total IGF-I, only marginally significant correlations were found between free IGF-I and birthweight (r = 0.25). According to multiple regression analysis free IGF-I levels were only dependent upon total IGF-I, IGFBP-2 and IGFBP-1, whereas IGFBP-3 levels did not contribute to the variance of free IGF-I concentrations in venous cord blood. There was no significant correlation between IGF-II and auxologic data at birth. When IGF-I and IGFBP-3 levels were analysed with respect to gestational age a biphasic pattern with maxima at 270 d was observed. IGFBP-2 exhibited a reversed pattern with a minimum at 265 d of gestation. In conclusion, these data suggest that IGF-I and the IGFBPs, but not IGF-II, play a role in the regulation of late fetal growth and development.

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/1.73 m2) 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.

OBJECTIVE The aims of this investigation were (a) to study the presence of immunoreactive forms of the acid‐labile subunit (ALS) in different human biological fluids, (b) to define the age dependence of serum ALS in normal children and adults and (c) to compare the regulation of ALS by GH or IGF‐I in children with GH deficiency (GHD) and GH receptor deficiency (GHRD) before and after 1 year of therapy with GH or IGF‐I, respectively. DESIGN AND PATIENTS Selected human biological fluids from different consenting volunteers and serum from 68 normal children and 5 adults were analysed. Four children diagnosed as GHD and 7 children diagnosed as GHRD were treated with recombinant human (rh) GH at a dosage of 0.05 mg/kg/day s.c. or rhIGF‐I at a dosage of 120 μg/kg twice daily s.c., respectively, for 12 months. MEASUREMENTS Immunoreactive forms of ALS were studied by Western immunoblot using a specific rabbit antiserum derived against synthetic human ALS and quantified by laser densitometry analysis. Serum from children with GHD or GHRD were sampled before and at 6 and 12 months of therapy; serum from these patients had been also assayed at baseline for determination of IGF‐I and IGF binding protein (IGFBP)‐3 by radioimmunoassay and immunoradiometric assay, respectively. RESULTS An immunoreactive 85 kDa doublet of ALS was detected in serum, plasma, follicular, peritoneal and synovial fluid, but not in urine, seminal plasma, amniotic or extra‐embryonic coelomic fluids. Assessment of serum from newborns to adults revealed an age dependence; the ALS doublet was low, but detectable, in newborns, increased during adolescence and remained constant in adulthood. ALS levels were significantly lower in GHD ( P = 0.02) and in GHRD children ( P = 0.001) than in age‐matched controls. Treatment with rhGH in GHD children produced a 2.7‐fold increase in serum ALS concentrations at 6 months of therapy ( P = 0.01), which was maintained after 1 year of treatment ( P = 0.006), leading to normalization of ALS concentrations. In contrast, administration of rhIGF‐I to GHRD children failed to increase and normalize serum ALS levels either at 6 or 12 months of therapy. CONCLUSIONS Immunoreactive forms of acid‐labile subunit are present in serum and plasma, as well as in follicular, peritoneal and synovial fluids, suggesting that acid‐labile subunit can either cross the capillary barrier or be secreted locally. Acid‐labile subunit concentrations are age‐dependent with a sharp increase during adolescence, and are reduced in GH deficient and GH receptor deficient children. While treatment with rhGH is able to increase and normalize acid‐labile subunit concentrations in GH deficient children, therapy with rhIGF‐I fails to increase serum acid‐labile subunit levels in GH receptor deficient patients. These data suggest that acid‐labile subunit is directly GH‐regulated, and that IGF‐I cannot increase acid‐labile subunit levels, as assessed by Western immunoblot.

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/1.73 m(2)) 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 nor 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 IGFr-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.

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.

IGFBPs play an important role in IGF biological actions bq 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 bu 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.

Abstract The insulin-like growth factors (IGF-I and IGF-II) and their binding proteins (IGFBPs) have been implicated in regulating fetal growth and development. The aim of this study was to determine whether fetal IGFs correlate with auxologic data at birth and/or gestational age. Venous cord blood was obtained from 138 healthy newborns immediately after birth and clinical data were recorded using a standardized data sheet. For the determination of IGF-I and IGF-II, IGFBP-blocked radioimmunoassays were used. A coated-tube immunoradiometric assay was applied for the measurement of free IGF-I. IGFBP-1, -2, and -3 were measured using specific radioimmunoassays. IGF-I levels were 61 ± 21 ng ml -1 , median 61 ng ml -1 , range 19-114ng ml -1 : IGF-II levels were 466 ± 80ng ml -1 , median 457 ng ml -1 , range 311–701 ng ml -1 ; free IGF-I levels were 2.4 ± 1.8ng ml -1 , median 1.8 ng ml -1 , range 0.4–7.8ng ml -1 . The concentration of IGFBP-1 was 144± 110 ng ml -1 , median 113 ng ml -1 , range 20–626 ng ml -1 ; that of IGFBP-2 was 1165 ± 455ng ml -1 , median 1119ng ml -1 , range 440–3466 ng ml -1 . IGFBP-3 levels were 1272 ± 280 ng ml -1 , median 1272ng ml -1 , range 600–1966 ng ml -1 . IGF-I levels correlated significantly with IGFBP-3 levels ( r = 0.71). birthweight ( r = 0.48) and birth length ( r = 0.37). There were significant inverse correlations between IGF-I and both IGFBP-I ( r = - 0.45) and IGFBP-2 ( r = - 0.62). Although free IGF-I levels correlated ( r = 0.71) with total IGF-I, only marginally significant correlations were found between free IGF-I and birthweight ( r = 0.25). According to multiple regression analysis free IGF-I levels were only dependent upon total IGF-I, IGFBP-2 and IGFBP-1, whereas IGFBP-3 levels did not contribute to the variance of free IGF-1 concentrations in venous cord blood. There was no significant correlation between IGF-II and auxologic data at birth. When IGF-I and IGFBP-3 levels were analysed with respect to gestational age a biphasic pattern with maxima at 270 d was observed. IGFBP-2 exhibited a reversed pattern with a minimum at 265 d of gestation. In conclusion, these data suggest that IGF-I and the IGFBPs, but not IGF-II, play a role in the regulation of late fetal growth and development.

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 (P < 0.0001), a minimal increase in TBK (P < 0.05), and a substantial decrease in Fat (P < 0.01) compared with baseline values. The loss of body fat continued to be significant (P < 0.01) in the GH/IGF-I group treatment at 12 weeks, whereas the increase in FFM was minimal (P < 0.05). 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.

Insulin-like growth factors (IGFs) and their specific regulatory binding proteins (IGFBPs) are postulated to play a key role in bone metabolism. To date, IGFBP-2 through -6 have been characterized in bone cell systems. In this study we focused on IGFBP-1. Primary cultures of normal human osteoblasts derived from trabecular bone (hOB cells) expressed low levels of IGFBP-1 messenger RNA (mRNA), as determined by Northern analyses. Treatment of hOB cells with 1 microM cortisol or 100 nM dexamethasone for 20 h stimulated IGFBP-1 mRNA expression 5-fold and increased levels of immunoassayable IGFBP-1 in the conditioned medium 3-fold. Estradiol and progesterone had no effect. IGFBP-1 expression was not observed in U-2, TE-85, or MG-63 human osteosarcoma cell lines or in normal human fibroblasts. Insulin (1-100 nM) potently inhibited both basal and glucocorticoid-stimulated IGFBP-1 expression in hOB cells. Insulin had little or no effect on steady state levels of the other IGFBP mRNA. A monoclonal antibody to the insulin receptor blocked insulin binding to insulin receptors and completely prevented insulin-induced suppression of IGFBP-1. In summary, we have documented IGFBP-1 mRNA and protein expression in normal nontransformed human osteoblastic cells. This expression was stimulated by glucocorticoids and inhibited by insulin in a manner similar to IGFBP-1 regulation in hepatocytes. Insulin acts through insulin receptors on hOB cells. We postulate that IGFBP-1 produced by osteoblasts in vivo can modulate local actions of IGF on bone formation in response to changes in glucocorticoid and insulin concentrations.