Relationships of serum insulin-like growth factor II concentrations to growth, compositional, and reproductive traits of swine

Insulin-like growth factors I and II (IGF-I and -II) are peptide hormones involved in metabolic regulation of growth. The objective of this study was to determine whether IGF-II concentration was predictive of growth, compositional, and reproductive traits of pigs. Forty male and sixty female pigs, divided equally between two locations, were weighed at 3-wk intervals from birth to 21 wk and bled at 9 and 21 wk of age. At each sampling, two blood samples were collected via jugular venipuncture at an interval of at least 1 h. Serum was separated and IGF-I, IGF-II, and growth hormone (GH) concentrations were determined via RIA. Traits measured included age at puberty and first parity litter size for gilts and backfat and longissimus muscle area. Blood was collected from a random sample of 52 progeny from 13 litters at 9 wk of age and serum was assayed for IGF-II concentrations. Effects of age, sex, location, and pig within sex x location on square-root transformed IGF-II concentrations were determined by analyzing data as a splitplot. Performance traits were fitted to a model including the effects of IGF-II concentration and combinations with IGF-I concentration, sex, location, and interactions. Concentrations of IGF-II were greater at 9 than at 21 wk of age (226.7 vs 159.3 ng/mL, respectively; P < .001) but did not differ between sexes. The correlation between serum IGF-II concentrations assayed from samples collected at 9 and 21 wk was .08. The partial correlations between IGF-I and IGF-II concentrations were .33 and .14 at 9 and 21 wk, respectively. The heritability of IGF-II concentration estimated from offspring-midparent regression was .08 +/- .20. Nine-week IGF-II concentration was positively associated with increased weight from weaning to 12 wk (P < .001). However, the sum of 9-wk IGF-I and IGF-II concentrations had a greater relationship to weight and gain in the growing phase than the concentration of either hormone alone. Concentration of IGF-II at 9 or 21 wk alone did not affect backfat thickness, longissimus muscle area, percentage lean, days to 100 kg, weight at 21 wk, age at puberty, or litter size. The sum of IGF-I and IGF-II concentrations was, however, associated with increased backfat and decreased days to 100 kg.     

Intramuscular collagen characteristics of ram, wether, and zeranol-implanted ram lambs

Eighteen spring-born Columbia ram, wether, and zeranol-implanted ram lambs were studied to determine the influence of castration or zeranol implants on intramuscular collagen (IMC) properties and muscle shear force values. Warner-Bratzler shear force values for longissimus muscle were greatest for ram lambs, intermediate for implanted rams, and least for wethers (P < .05). Nonreducible collagen crosslink concentration was greater in IMC of rams and implanted rams (P < .05). The IMC from rams compared with that from wethers contained proportionately more Type III than Type I collagen (P < .05); values for implanted rams were intermediate. Heat-soluble muscle collagen concentration was greater for rams and implanted rams than for wethers (P < .05); however, insoluble collagen concentration did not differ by treatment. Muscle collagen concentrations were not different for rams, wethers, or implanted rams. Increased shear force values in rams were associated with elevated collagen crosslink concentration and increased proportion of Type III collagen. Greater concentration of soluble collagen in ram IMC neither diminished nor diluted IMC crosslinking. The proportion of heat-labile collagen in the fractions did not reflect the IMC crosslinking profile for ram and wether lambs. Zeranol implantation modified IMC characteristics of rams such that shear force values and some collagen properties were similar to those of wethers.     

Effect of testosterone on differential muscle growth and on protein and nucleic acid concentrations in muscles of growing lambs

Growth, nucleic acid, and protein concentrations were measured in three muscles of 20 rams, 20 wethers, and 20 wethers implanted with testosterone. Two lambs from each group were slaughtered at 14-d intervals from 49 to 133 d, and then at 28-d intervals until 217 d, for a total of 10 slaughter ages. Immediately after slaughter, the semitendinosus, splenius, and triceps brachii muscles were removed, trimmed of adhering fat, and weighed. The DNA, RNA, and protein concentrations of these muscles were determined. Testosterone increased combined weight of the three muscles. The splenius muscles of rams and wethers implanted with testosterone were heavier and had a biphasic growth pattern as the combined muscle weight increased, whereas the splenius muscle of wethers had a single growth phase. Rams and implanted wethers had greater splenius muscle DNA and RNA concentrations than wethers as muscle weight increased. This model could be used to study the gene regulation of testosterone-induced muscle growth with the possibility of invoking similar effects in more economically important muscles.     

Pituitary and plasma levels of growth hormone in Booroola sheep that are either homozygous carriers or non-carriers of the FecBB fecundity gene

The aim of this study was to examine the effect of the FecBB fecundity gene on plasma concentrations and pituitary content of growth hormone (GH) in sheep. No differences were found between homozygous carriers (BB) and non carriers (++) of the FecBB gene with regard to pituitary GH contents in both ovariectomized and intact ewes. However, ovariectomized ewes had higher levels of pituitary GH than intact ewes (P < 0.01). There were no differences between FecBB genotypes with respect to plasma concentrations of GH in 6-year-old ovariectomized ewes bled every 10 min for 12 h or in ram lambs bled weekly during their first year of life. GH levels in the rams decreased until week 27, increased to a peak at week 31 then decreased before increasing again at week 43. Mean plasma GH concentrations in the ewe lambs bled weekly for a year decreased until week 19 then remained at approximately this level for the remainder of the year. Mean GH plasma concentrations in the ram lambs were higher than in the ewe lambs (P < 0.001). Ewe lambs that were homozygous for the FecBB gene had lower body weights (P < 0.05) and had higher levels of GH (P < 0.01) than non carrier ewe lambs during their first year. Before the average age of first behavioural oestrus (36 weeks) GH levels in the ewe lambs were negatively correlated with body weights (r = -0.69, P < 0.001, n = 22). When body weight was included as a covariate in analysis of variance the genotype difference in ewe lamb plasma GH concentrations was no longer significant.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effects of treatment with megestrol acetate, aminoglutethimide, or formestane on insulin-like growth factor (IGF) I and II, IGF-binding proteins (IGFBPs), and IGFBP-3 protease status in patients with advanced breast cancer

The effects of treatment with the aromatase inhibitors aminoglutethimide (AG) and formestane or the synthetic progestin megestrol acetate (MA) on plasma levels of insulin-like growth factor I (IGF-1), IGF-II, IGF-binding proteins (IGFBPs), and IGFBP-3 protease status were investigated in 39 patients suffering from advanced breast cancer. Treatment with AG and MA elevated plasma levels of IGF-I by mean values of 27% (n = 15; P < 0.025) and 81% (n = 7; P < 0.025), respectively, whereas treatment with formestane had no effect (n = 13). Treatment with AG increased plasma levels of IGFBP-2, as evaluated by Western blotting (P < 0.01). MA caused a significant reduction in IGFBP-3 protease activity (mean reduction, 69%; P < 0.05). These alterations in plasma IGF-I and IGFBP-3 protease activity were reversed 4 weeks after terminating MA therapy (n = 8; P < 0.025). Taken together, 13 of 15 patients had reduced IGFBP-3 protease activity during treatment with MA compared to the control situation (P < 0.0025). Total levels of IGFBP-3 as measured by RIA were moderately elevated by treatment with MA (mean increase, 19%; P < 0.05), and Western immunoblotting revealed an increase in the amount of intact IGFBP-3 and reduced amounts of IGFBP-3 in the modified form. None of the treatment modalities had any influence on plasma levels of IGF-II. The increase in the plasma IGF-I concentration seen during treatment with MA may be secondary to an increased level of intact IGFBP-3. This could reflect an alteration in IGF availability that contributes to the antitumor effect of MA.     

Developmental pattern of fetal growth hormone, insulin-like growth factor I, growth hormone binding protein and insulin-like growth factor binding protein-3

AIMS: To evaluate the developmental pattern of fetal growth hormone (GH), insulin-like growth factor I (IGF-I), GH binding protein (GHBP) and IGF binding protein-3 (IGF-3); to determine the implications for fetal growth. METHODS: Serum GH, IGF-I, GHBP and IGFBP-3 were measured in 53 fetuses, 41 aged 20-26 weeks (group A) and 12 aged 31-38 weeks (group B). Fetal blood samples were obtained by direct puncture of the umbilical vein in utero. Fetal blood samples were taken to rule out beta thalassaemia, chromosome alterations, mother to fetus transmissible infections, and for maternal rhesus factor. GHBP was determined by gel filtration chromatography of serum incubated overnight with 125I-GH. GH, IGF-I and IGFBP-3 were determined by radioimmunoassay. RESULTS: Fetal serum GH concentrations in group A (median 29 micrograms/l, range 11-92) were significantly higher (P < 0.01) than those of group B (median 16.7 micrograms/l, range 4.5-29). IGF-I in group A (median 20 micrograms/l, range 4.1-53.3) was significantly lower (P < 0.01) than in group B (median 75.2 micrograms/l, range 27.8-122.3). Similarly, IGFBP-3 concentrations in group A (median 950 micrograms/l, range 580-1260) were significantly lower than those of group B (median 1920 micrograms/l, range 1070-1770). There was no significant difference between GHBP values in group A (median 8.6%, range 6.6-12.6) and group B (median 8.3%, range 6-14.3). Gestational age correlated positively with IGF-I concentrations (P < 0.0001) and IGFBP-3 (P < 0.0001) and negatively with GH (P < 0.0001). GHBP values did not correlate with gestational age. Multiple regression analysis showed a negative correlation between GH:IGF-I ratio and fetal growth indices CONCLUSIONS: The simultaneous evaluation of fetal GH, IGF-I, IGFBP-3 and GHBP suggests that the GH-IGF-I axis might already be functional in utero. The progressive improvement in the efficiency of this axis in the last part of gestation does not seem to be due to an increase in GH receptors.     

Role for membrane and secreted insulin-like growth factor-binding protein-2 in the regulation of insulin-like growth factor action in lung tumors

The insulin-like growth factors (IGFs) have been implicated in the autocrine and/or paracrine growth of a number of tumor types, including lung tumors. Importantly, insulin-like growth factor-binding proteins (IGFBPs), which both enhance and inhibit the physiological and biological actions of the IGFs, have been shown to be secreted in vitro by a wide range of tumors. In particular, IGFBP-2 is frequently produced by human tumor cells, suggesting that this protein may be an important determinant of IGF action in tumors. In the present study, we investigated IGFBP-2 effects in lung tumor cells by examining the influence of IGFBP-2 on IGF-receptor interaction and the biological actions of IGF-I and IGF-II. Affinity cross-linking studies demonstrated expression of type-I and type-II IGF receptors on small cell lung cancer (SCLC) and non-small cell lung cancer (NSCLC) cells and the presence of abundant membrane-associated IGFBP in SCLC cells but not in NSCLC cells. An antiserum specific for IGFBP-2 was used in immunoprecipitation and immunoblotting studies which demonstrated that the membrane-associated IGFBP identified by affinity cross-linking in SCLC cells is IGFBP-2. In NSCLC cells, both IGF-I and IGF-II bound predominantly to IGF-I receptors, whereas in SCLC cells binding was principally to surface-associated IGFBP-2. SCLC cells failed to respond to IGF-I and -II stimulation in a DNA synthesis assay. For NSCLC cells, IGF-II was a more potent stimulator of DNA synthesis than IGF-I. Soluble IGFBP-2 inhibited the binding of radiolabeled IGF-I and -II to both SCLC and NSCLC cells in a concentration-dependent manner and inhibited IGF-stimulated DNA synthesis in NSCLC cells. These observations indicate that both soluble and membrane-associated IGFBP-2 compete with IGF receptors for ligand binding and, thus, are likely to be important determinants of IGF responsiveness. The findings of the present study also indicate that the type-I receptor on NSCLC cells contains a high-affinity binding site for IGF-II which presumably mediates the biological effects of IGF-II in these cells, thereby implicating IGF-II in the autocrine/paracrine growth of NSCLC.     

Long-term effects of insulin-like growth factor (IGF)-I treatment on serum IGFs and IGF binding proteins in adolescent patients with growth hormone receptor deficiency

OBJECTIVE: The aim of this investigation was to study the effect of relatively high dose IGF-I therapy given for several months, on serum levels of IGF-I, IGF-II and IGFBP-3, and on IGF-I pharmacokinetics in patients with growth hormone insensitivity due to GH receptor dysfunction. DESIGN AND PATIENTS: Two adolescent subjects from Ecuador were treated with recombinant IGF-I at a dosage of 120 micrograms/kg s.c. twice daily, in combination with a GnRH analogue for 8 months. MEASUREMENTS: Serum was sampled at baseline and at 3-8 months, for determination of IGF-I, IGF-II and IGFBP-3 by radioimmunoassay, and for evaluation of IGFBPs and IGFBP-3 protease activity by Western ligand blot and protease assay, respectively. RESULTS: Peak serum IGF-I levels ranged from 272 to 492 micrograms/l. Mean serum IGF-II levels were decreased concurrently with the increase in IGF-I. Serum IGFBP-3 levels failed to rise with prolonged IGF-I treatment. There was no apparent change in the half-life of IGF-I during the treatment period. CONCLUSIONS: IGF-I administration does not increase serum levels of IGFBP-3 or significantly alter IGF-I pharmacokinetics.     

Insulin-like growth factor I in fetal serum obtained by cordocentesis is correlated with intrauterine growth retardation

We examined whether insulin-like growth factor-I (IGF-I) and one of its binding proteins (IGFBP-1) in fetal serum obtained by cordocentesis is correlated with intrauterine growth retardation (IUGR) and weight estimation by ultrasound. Cordocentesis sera from 27 fetuses suspected of having IUGR were analysed for IGF-I and IGFBP-1 by radioimmunoassay. The results showed that IGF-I concentrations were correlated significantly with birth weight (P < 0.001) and placenta weight (P < 0.05). Mean fetal concentrations of IGF-I were 38 +/- 18 microg/l. In patients (n = 11) with a weight deviation at delivery <-33%, IGF-I concentrations were 24.1 +/- 13.2 microg/l. IGFBP-1 was inversely correlated with birth weight (P < 0.006) and concentrations of IGF-I. Mean plasma concentrations of IGFBP-1 were 234.2 +/- 161.4 microg/l. Furthermore, IGF-I concentrations were correlated with the weight deviation estimated by ultrasonography at the time of cordocentesis (P < 0.007), as well as with the weight deviation at delivery (P < 0.0001). The actual weight deviation at delivery was correlated more strongly with fetal IGF-I concentrations than with the estimated weight deviation at cordocentesis. The lowest concentrations of IGF-I were found in patients with a weight deviation <-33%. Very low concentrations of IGF-I are thus associated with IUGR, indicating that IGF-I measured in fetal serum may increase the predictive value of ultrasonographic weight estimation.     

Free and total insulin-like growth factors and insulin-like growth factor binding proteins during 14 days of growth hormone administration in healthy adults

The objective was to investigate the effect of growth hormone (GH) administration on circulating levels of free insulin-like growth factors (IGFs) in healthy adults. Eight healthy male subjects were given placebo and two doses of GH (3 and 6 IU/m2 per day) for 14 days in a double-blind crossover study. Fasting blood samples were obtained every second day. Free IGF-I and IGF-II were determined by ultrafiltration of serum. Total IGF-I and IGF-II were measured after acid-ethanol extraction. In addition, GH, insulin, IGF binding protein 1 (IGFBP-1) and IGFBP-3 were measured. Serum-free and total IGF-I increased in a dose-dependent manner during the 14 days of GH administration. After 14 days, serum-free IGF-I values were 610 +/- 100 ng/l (mean +/- SEM) (placebo), 2760 +/- 190 ng/l (3 IU/ m2) and 3720 +/- 240 ng/l (6 IU/m2) (p = 0.0001 for 3 and 6 IU/m2 vs placebo; p = 0.004 for 3 IU/m2 vs 6 IU/m2). Total IGF-I values were 190 +/- 10 micrograms/l (placebo), 525 +/- 10 (3 IU/m2), and 655 +/- 40 micrograms/l (6 IU/m2) (p < 0.0001 for 3 and 6 IU/m2 vs placebo; p = 0.04 for 3 IU/m2). There were no differences in the levels of free or total IGF-II during the three study periods. Insulin-like growth factor binding protein 1 was decreased during GH administration (p = 0.04 for placebo vs 3 IU/m2; p = 0.006 for placebo vs 6 IU/m2). In conclusion, fasting serum free IGF-I increased dose dependently during GH administration and free IGF-I increased relatively more than total IGF-I. This may partly be due to the decrease in IGFBP-1.     

Free and total insulin-like growth factor I (IGF-I), IGF-binding protein-1 (IGFBP-1), and IGFBP-3 and their relationships to the presence of diabetic retinopathy and glomerular hyperfiltration in insulin-dependent diabetes mellitus

The existing literature on serum insulin-like growth factor I (IGF-I) levels in insulin-dependent diabetes mellitus (IDDM) is conflicting. Free IGF-I may have greater physiological and clinical relevance than total IGF-I. Recently, a validated method has been developed to measure free IGF-I levels in the circulation. Serum free and total IGF-I, IGF-binding protein-1 (IGFBP-1), and IGFBP-3 levels were measured in 56 insulin-treated IDDM patients and 52 healthy sex- and age-matched controls. Diabetic retinopathy was established by direct fundoscopy. In 54 IDDM patients, the glomerular filtration rate (GFR) and effective renal plasma flow were calculated from the clearance rate of [125I]iothalamate and [131I]iodohippurate sodium. Fasting free IGF-I, total IGF-I, and IGFBP-3 levels were significantly lower in IDDM patients than in age- and sex-matched healthy controls (free IGF-I, P < 0.005; total IGF-I, P < 0.001; IGFBP-3, P = 0.001), whereas IGFBP-1 levels were higher (P < 0.001). In IDDM subjects, decreases in free IGF-I, total IGF-I, and IGFBP-3 levels with age were observed (free IGF-I, r = -0.27 and P = 0.05; total IGF-I, r = -0.52 and P < 0.001; IGFBP-3, r = -0.37 and P = 0.005). Free IGF-I was inversely related to fasting glucose in IDDM subjects (r = -0.35; P = 0.01), whereas the relationship between total IGF-I and fasting glucose did not reach significance (r = -0.27; P = 0.06). Age-adjusted free IGF-I levels were significantly higher (P < 0.05) in IDDM subjects with retinopathy than in subjects without retinopathy after adjustment for age. Total IGF-I and IGFBP-3 levels were positively related to GFR (total IGF-I, r = 0.35 and P < 0.05; IGFBP-3, r = 0.28 and P < 0.05). Both of these differences lost significance after adjustment for age. Free IGF-I, total IGF-I, and IGFBP-3 levels were lower and IGFBP-1 levels were higher in insulin-treated IDDM subjects compared to those in age- and sex-matched controls. Free IGF-I, total IGF-I, and IGFBP-3 levels decreased significantly with age in IDDM subjects. Age-adjusted free IGF-I levels in subjects with diabetic retinopathy were higher than those in subjects without diabetic retinopathy. Total IGF-I and IGFBP-3 levels were positively related to GFR in IDDM subjects, but these relations were lost after adjustment for age. Measurement of serum free IGF-I levels in IDDM subjects did not have clear advantages compared to that of total IGF-I, IGFBP-1, and IGFBP-3 levels. Serum IGF-I and IGFBPs reflect their tissue concentrations to a various degree. Consequently, extrapolations concerning the pathogenetic role of the IGF/IGFBP system in the development of diabetic complications at the tissue level remain speculative.     

Serum insulin-like growth factor-I, insulin-like growth factor binding protein-3, sex steroids, osteocalcin and bone mineral density in male and female rats

Although it has been reported that the rate of weight gain and linear growth increases markedly during puberty in rats, little is known about the relationship between endocrine changes and bone mineral density (BMD) changes upon sexual maturation in these animals. The aim of this study was to examine the levels of serum insulin-like growth factor-I (IGF-I), IGF binding protein (IGFBP)-3, sex steroids and osteocalcin, and the changes in BMD in normal aging male and female rats. Male rats exhibited increases in serum IGF-I and IGFBP-3 concentrations before increases in serum testosterone levels. IGF-I and testosterone peaked at 9 weeks of age, and thereafter remained in a steady state, whereas IGFBP-3 reached a peak at 7 weeks of age, and then gradually declined. A strong correlation between serum IGF-I and IGFBP-3 levels was found in subjects 3-9 weeks old. A highly significant correlation between serum IGF-I and testosterone levels was also found. In females, serum 17 beta-estradiol, IGF-I and IGFBP-3 levels increased gradually from 3 to 5 weeks old, peaked at 9 weeks, and then decreased slowly thereafter. The correlation coefficient between serum IGF-I and IGFBP-3 was highly significant. The correlation coefficient between serum IGF-I and 17 beta-estradiol levels was weak, although it was strongest when the subjects were 3-9 weeks old. Serum osteocalcin is a marker of bone formation; its level remained relatively high from 3 to 9 and from 3 to 7 weeks of age in males and females, respectively, although osteocalcin in both sexes declined gradually with age. As for bone mass, sharp increases in BMD in the tibia, femur and lumbar vertebrae appeared earlier in female than in male rats, and the BMD in females tended to be higher than in males between 5 and 9 weeks old. After 9 weeks of age, BMD in males was higher than that in females, as BMD in males continued to increase whereas females tended to remain in a steady state after this stage. The correlation coefficients between tibial BMD and serum IGF-I or IGFBP-3 levels were highly significant when the subjects were from 3 to 9 weeks old. Taken together, these results suggest that BMD development occurs earlier in female than in male rats. This sex-related difference in changes in the BMD pattern may result from the earlier onset of puberty in females, and from sex-specific differences in concentrations of IGF-I, IGFBP-3 and sex steroids during maturation.     

Increases in insulin-like growth factor binding protein-2 accompany decreases in proliferation and differentiation when porcine muscle satellite cells undergo multiple passages

Subjecting cloned porcine myogenic satellite cells to multiple passages leads to decreased rates of cell division and myotube formation. Because IGF have been implicated in the regulation of muscle cell proliferation and differentiation, the present study was conducted to characterize secretion of IGF-I and IGF-binding proteins (IGFBP) in cultures of cloned porcine satellite cells at two stages of multiple passaging. To this end, we obtained a single porcine satellite cell clone that demonstrated relatively high capacities for cellular proliferation and differentiation into myotubes at the fifth passage but that had greatly diminished capacities for proliferation and myotube formation by the seventh passage. The predominant IGFBP secreted by this satellite cell clone was immunologically identified as IGFBP-2, and quantities of it were increased in medium from seventh-passage cultures. Quantities of IGF-I in medium were determined with a newly developed "titration" radioimmunoassay in which interference from IGFBP was minimized by adding a range of saturating quantities of IGF-II. Medium IGF-I concentrations in seventh-passage cultures were also increased relative to the fifth-passage cultures when expressed per unit of DNA. It is hypothesized that the observed increase of IGF-I in medium likely resulted from protective sequestration of IGF-I by IGFBP-2 rather than from enhanced IGF-I secretion. In summary, these data suggest that multiple passaging of cloned porcine satellite cells results in increased secretion of IGFBP-2, which is associated with depressed cell proliferation and myotube formation, perhaps because the increased IGFBP-2 sequestered IGF-I and reduced its bioactivity.     

Ontogeny of insulin-like growth factors (IGF), IGF binding proteins, IGF receptors, and growth hormone receptor mRNA levels in porcine pancreas

We examined the ontogeny of mRNA levels of IGF-I and -II, IGF type 1 (IGFI-R) and type II receptors (IGFII-R), IGF binding protein-1 and -3 (IGFBP-1 and -3), GH receptor (GHR), and tissue concentrations of IGF and IGFBP in the pancreas of pigs. Tissues were collected from fetuses at 90 and 110 d of gestation and from pigs at 1, 21, 90 and 180 d of age. Northern blots were performed using total RNA hybridized with 32P-labeled cDNA probes (human IGF-I and human IGFI-R) and cRNA probes (rat IGF-II, human IGFII-R, human IGFBP-1, pig IGFBP-3, and pig GHR). There were two accelerated growth stages of the pancreas: the first one at 90 d of fetal life, which is characterized by cell hyperplasia (high ratio of DNA to body weight), and the second one at postnatal 90 d, which is attributed to cell hypertrophy (high ratios of pancreatic weight, RNA, and protein to DNA). The level of IGF-II mRNA and its tissue concentration were predominant during fetal life and low thereafter. The IGF-I mRNA level was high during fetal and early postnatal life and decreased thereafter. Messenger RNA levels of IGFI-R, IGFBP-3, and GHR and concentrations of IGFBP-1 and -2 were abundant during fetal and early postnatal life. In conclusion, IGF may be involved in various physiological periods of pancreatic development in pigs.     

Follicular fluid insulin-like growth factor-I and insulin-like growth factor-binding protein-1 and -3 vary as a function of ovarian reserve and ovarian stimulation

PURPOSE: Follicular fluid concentrations of insulin-like growth factor (IGF)-I, IGF-II, IGF-binding protein (BP)-1, and IGFBP-3 in 57 women undergoing in vitro fertilization and embryo transfer were examined to determine whether levels reflected differences in patients' exposure to gonadotropin stimulation and a diminished ovarian reserve. METHODS: Preovulatory follicular fluid was obtained from both gonadotropin-stimulated and unstimulated cycles. Subjects were grouped according to normal or decreased ovarian reserve and whether or not they received gonadotropin stimulation. RESULTS: The mean follicular fluid concentrations of IGF-I and IGFBP-1 were significantly lower in the "decreased" ovarian reserve group compared with the "normal" ovarian reserve group, with no change in estradiol or IGF-II levels. This resulted in a decreased molar IGF-I: BP ratio and an increased molar IGF-II:IGFBP-1 ratio. In unstimulated cycles, mean follicular fluid concentrations of IGFs did not differ significantly compared with those in stimulated cycles, whereas concentrations of IGFBP-1 and IGFBP-3 were significantly lower, leading to higher molar ratios of the IGFs to the binding proteins. CONCLUSIONS: Follicular fluid IGF and binding proteins vary as a function of ovarian reserve and gonadotropin stimulation. This may reflect either differences in oocyte quality or a suboptimal follicular fluid environment.     

Insulin-like growth factor binding proteins-2 and -3 stimulate growth hormone receptor binding and mitogenesis in rat osteosarcoma cells

GH exerts its biological actions on osteoblasts through a specific high affinity receptor expressed on these cells. GH receptor binding is positively modulated by a number of factors, including retinoic acid and dexamethasone, whereas fetal calf serum strongly decreases the binding. To identify responsible factors in serum, components of serum, the insulin-like growth factors (IGFs)-I and -II, and IGF binding proteins (IGFBPs)-2 and -3 were tested for a possible negative modulatory role. IGF-I and -II decreased [125I]hGH binding at an optimal concentration of 30 ng/ml for IGF-I and 100 ng/ml IGF-II, reducing the binding to 51% and 55%, respectively, of control values. A stimulation of [125I]hGH binding was observed with IGFBP-2 as well as IGFBP-3, inducing an increase to 148% and 151% of control binding at an optimal concentration of 3000 ng/ml for both peptides. The effects of all peptides were dependent on the incubation time, being significantly increased after 8 h of incubation and reaching the full effect thereafter. The effects were declined at 24 h compared with 16 h for IGFBP-2 and -3 but not for IGF-I and -II. Coincubation of the cells with IGF-I and -II and IGFBP-2 and -3 neutralized the effects of the factors alone. In conclusion, these results show that IGF-I and -II on the one hand and IGFBP-2 and -3 on the other hand exert opposite actions on [125I]hGH binding, IGFBP-2 and -3 exerting probably an IGF-independent effect. Further, IGF-I and -II decreased GH receptor messenger RNA (mRNA) levels, as quantified by a solution hybridization ribonuclease protection assay, from 8.65 +/- 1.78 attomoles (amol)/microgram DNA (control) to 2.4 +/- 0.68 and 2.16 +/- 0.92 amol/microgram DNA, respectively. IGFBP-2 increased GH receptor mRNA levels from 5.26 +/- 1.17 (control) to 13.19 +/- 3.48. Incubation with IGFBP-3 did not result in stimulation of GH receptor mRNA levels (8.59 +/- 2.91 amol/microgram DNA). This shows that the mechanism of regulation of the GH receptor is, except for IGFBP-3, at least in part on the mRNA level. Lastly, IGFBP-2 and IGFBP-3 are mitogenic for UMR-106.01 rat osteosarcoma cells, inducing an increase in cell number to 125% and 142% of control cell counts after 48 h of incubation with 1000 ng/ml IGFBP-2 and -3, whereas IGF-I, IGF-II and Long R3 IGF-I did not stimulate proliferation. IGFBP-2 and -3 potentiate hGH induced mitogenesis at low hGH concentrations of both factors, whereas at higher concentrations no such effect is observed.(ABSTRACT TRUNCATED AT 400 WORDS)     

Acute effects of short-term feed deprivation and refeeding on circulating concentrations of metabolites, insulin-like growth factor I, insulin-like growth factor binding proteins, somatotropin, and thyroid hormones in adult geldings

Two studies were performed with Standardbred geldings 7 to 21 yr of age to determine the sequence of changes in blood plasma concentrations of some hormones and metabolites during feed deprivation for 48 h and for 12 h after refeeding. Plasma hormone and metabolite concentrations were determined with methods validated for horse plasma. Insulin-like growth factor binding proteins (IGFBP) were determined with radioligand analysis following SDS-PAGE electrophoresis. In both experiments, plasma concentrations of triiodothyronine and thyroxine decreased (P < .01) during feed deprivation and increased (P < .01) during refeeding. Plasma glucose and IGF-I either decreased or were not altered during feed deprivation. In contrast, plasma concentrations of NEFA and urea nitrogen increased (P < .01) during feed deprivation and decreased (P < .01) during the refeeding period. Plasma somatotropin (ST) increased (P < .01) approximately 80% at 24 to 36 h of feed deprivation, declined (P < .01) to control values at 48 h of feed deprivation, increased (P < .01) nearly three fold at 3 h after refeeding, and returned to control values by 6 h after refeeding. We identified five IGFBP, and their plasma concentrations were not significantly altered during feed deprivation or following refeeding. We conclude that metabolite availability during feed deprivation and following refeeding alters the secretion of thyroid hormones, ST, and possibly IGF-I, thereby maintaining homeostasis in horses.     

Porcine insulin-like growth factor (IGF)-binding protein-3 elicits bi-phasic effects on IGF-I stimulated DNA synthesis in neonatal porcine skin fibroblasts

The present study was undertaken to investigate the effects of porcine IGFBP-3 on IGF-I stimulated DNA synthesis in neonatal porcine skin fibroblasts. IGF-I stimulated DNA synthesis in skin fibroblasts in a concentration dependent manner. DNA synthesis was maximally stimulated by 5 to 20 fold at 5 nM IGF-I; half-maximal stimulation was observed at approximately 1 nM IGF-I. Co-incubation of IGFBP-3 with a maximally effective dose of IGF-I (10 nM) did not inhibit the stimulatory effects of IGF-I on DNA synthesis. In contrast, when IGFBP-3 at concentrations of 0 to 20 nM was co-incubated with 1 nM IGF-I, a bi-phasic dose response was observed with IGFBP-3 being inhibitory only at a 10 to 20 fold molar excess to IGF-I. Based on the approximately equal molar ratio of IGFBP-3:IGF-I present in the circulation of control and pST-treated pigs our results suggest that IGFBP-3 does not inhibit the mitogenic effects of IGF-I. In summary, these results indicate that the combination of IGFBP-3 with IGF-I optimizes mitogenic signalling via the type I IGF receptor and suggest that IGFBP-3 does not inhibit the effects of ST that are mediated by IGF-I.     

Filter paper blood spot assay of human insulin-like growth factor I (IGF-I) and IGF-binding protein-3 and preliminary application in the evaluation of growth hormone status

To facilitate broader applications of insulin-like growth factor I (IGF-I) and IGF-binding protein-3 (IGFBP-3) analysis, we developed procedures for their measurements in extracts of whole blood dried on filter paper. A single 8-mm diameter filter paper disc containing about 13 microL blood was used. IGFBP-3 was efficiently extracted in a buffer within 1 h of incubation. IGF-I extraction involved incubation in buffer followed by acidification and neutralization steps. Blood spot assays showed intra- and interassay coefficients of variation (including interspot variations) of 5.4-16.7% for IGF-I and 6.6-11.7% for IGFBP-3; recoveries were 97 +/- 7.1% and 101 +/- 8.7%, respectively. Recoveries of IGF-I and IGFBP-3 in response to 4- to 8-fold variations in extraction buffer volume were 97 +/- 8.2% and 107 +/- 6.1%, respectively. Dried blood spot IGF-I and IGFBP-3 showed greater than 1-month stability at -20 C, 4 C, and room temperature and retained more than 65% of the immunoreactivity after approximately 1 month at 37 C. Both IGF-I and IGFBP-3 were contained within the plasma fraction of whole blood, and variations (mean +/- SD) in IGF-I (204 +/- 29 micrograms/L) and IGFBP-3 (4.4 +/- 0.48 mg/L) measured in extracts of dried blood spot with adjusted hematocrit of 0.2-0.62 were acceptable. IGF-I and IGFBP-3 in paired plasma and dried blood spot extracts of random samples (n = 46) showed excellent correlation (r > 0.94) with slopes of near unity. Compared to conventional methods, the filter paper procedures were equally effective in distinguishing IGF-I and IGFBP-3 levels in untreated GH receptor-deficient (n = 11) and age-matched normal controls (n = 16). We conclude that blood collected on filter paper is ideal for IGF-I and IGFBP-3 analysis and may find applications in pediatric and large scale infant screening programs.