Insulin-like growth factors I and II. Peptide, messenger ribonucleic acid and gene structures, serum, and tissue concentrations.

There is currently widespread interest in the IGFs (IGF-I and IGF-II) and their roles in the regulation of growth and differentiation of an ever increasing number of tissues are being reported. This selective review focused on the current state of our knowledge about the structure of mammalian IGFs and the multiple forms of mRNAs which arise from alternative splicing and promoter sites which arise from gene transcription. Current progress in the immunological measurement of the IGF is reviewed including different strategies for avoiding binding protein interference. The results of measurements of serum IGF-I and IGF-II in fetus and mother and at various stages of postnatal life are described. Existing knowledge of the concentration of these peptides in body fluids and tissues are considered. Last, an attempt is made to indicate circumstances in which the IGFs are exerting their actions in an autocrine/paracrine mode and when endocrine actions predominate. In the latter context it was concluded that an important role for GH action on skeletal tissues via hepatic production of IGF-I and endocrine action of IGF-I on growth cartilage is likely.

Fasting enhances growth hormone secretion and amplifies the complex rhythms of growth hormone secretion in man.

Studies in man have shown that the episodic release of growth hormone (GH) is infrequent and erratic, and unlike that in the rat does not appear to have discernible ultradian periodicities. However, these observations in nonfasted subjects may be invalid since mixed nutrients have unpredictable effects on GH release. Moreover, in the fed state basal GH levels are frequently undetectable, thus rendering the identification of low amplitude pulses unreliable. Accordingly, the 24-h pulsatile pattern of GH secretion obtained from repetitive venous sampling in six normal adult male subjects was examined during a control fed day and during the first and fifth days of a 5-d fast. The GH data were analyzed using two distinct methods: a discrete pulse detection algorithm (Cluster analysis) and Fourier expansion time-series, which allows fixed periodicities of secretory activity to be resolved. The 5-d fast resulted in a significant increase in discrete GH pulse frequency (5.8 +/- 0.7 vs. 9.9 +/- 0.7 pulses/24 h, P = 0.028), 24 h integrated GH concentration (2.82 +/- 0.50 vs. 8.75 +/- 0.82 micrograms.min/ml; P = 0.0002), and maximal pulse amplitude (5.9 +/- 1.1 vs. 12.3 +/- 1.6 ng/ml, P less than 0.005). While multiple low-amplitude sinusoidal periodicities were present on the control fed day, time-series analysis revealed enhancement of circadian and ultradian cycles on the first and fifth days of fasting. Concomitantly, fasting resulted in a decline (day 1 vs. day 5) in serum concentrations of somatomedin C (1.31 +/- 0.22 vs. 0.77 +/- 0.18 U/ml) and glucose (4.9 +/- 0.2 vs. 3.2 +/- 0.2 mmol/liter), and a marked rise in free fatty acid (0.43 +/- 0.12 vs. 1.55 +/- 0.35 mmol/liter) and acetoacetate (35 +/- 6 vs. 507 +/- 80 nmol/liter). We conclude that the acute nutritional status is an important determinant of spontaneous pulsatile GH secretion in man. Fast-induced enhancement of GH release is achieved through combined frequency (discrete pulses) and amplitude (sinusoidal periodicities) modulation. Such alterations in somatotropic hormone release may play an important role in substrate homeostasis during starvation.

Effect of tamoxifen on plasma insulin-like growth factor I in patients with breast cancer.

Human breast cancer cells secrete and have membrane receptors for insulin-like growth factor I (IGF-I), a growth hormone-dependent peptide that stimulates cell replication. However, little is known about plasma concentrations of IGF-I in breast cancer patients. Plasma IGF-I levels are decreased in malnutrition, decline with advancing age, and are influenced by estrogen. We evaluated the effect of the antiestrogen agent tamoxifen on plasma IGF-I in 32 ambulatory breast cancer patients. Treatment with tamoxifen was associated with lower concentrations of plasma IGF-I (0.48 +/- 0.3 unit/ml in treated versus 1.03 +/- 0.6 units/ml in nontreated patients, P less than 0.01). However, patients treated with tamoxifen did not differ from nontreated patients in age, menopause, duration since diagnosis, metastatic disease, recent weight loss, or measures of nutritional status. We conclude that tamoxifen therapy results in a reduction of plasma IGF-I concentration. We speculate that the antitumor action of tamoxifen in breast cancer is due in part to suppression of IGF-I.

Caloric restriction and 7,12-dimethylbenz(a)anthracene-induced mammary tumor growth in rats: alterations in circulating insulin, insulin-like growth factors I and II, and epidermal growth factor.

Caloric restriction (CR) inhibits many neoplastic diseases in rodents, yet the biochemical mechanism(s) for these effects are poorly understood. We have examined the effects of ad libitum (AL) feeding with 25 or 40% CR on the promotion of 7,12-dimethylbenz(a)anthracene-induced mammary tumorigenesis in virgin female Sprague-Dawley rats. Further, we have also studied the influence of chronic CR on temporal alterations in circulating insulin, insulin-like growth factor I/somatomedin C, insulin-like growth factor II/multiplication-stimulating activity, and epidermal growth factor levels at 0, 1, 3, 5, 11, and 20 weeks in carcinogen- and vehicle-treated animals. Tumor incidence and multiplicity were markedly inhibited (P less than 0.05) with increasing CR. Fasting serum insulin-like growth factor I/somatomedin C levels exhibited a significant acute decline with CR at 1 and 3 weeks, but were comparable to AL-fed controls throughout the remainder of the 5-month study, despite continued differences in weight gain between AL and CR rats. Levels of insulin-like growth factor II/multiplication-stimulating activity exhibited no discernible pattern in relation to CR. Serum insulin levels showed age-dependent increases, but were affected by increasing CR at all time points. Insulin levels were significantly (P less than 0.05) reduced in 40% CR rats from 3 weeks onward compared to controls, while 25% CR resulted in nonsignificant (P less than 0.07) reductions throughout the study. No significant differences in growth factor levels were observed between 7,12-dimethylbenz(a)anthracene- and vehicle-treated rats. Circulating epidermal growth factor was not detectable in any treatment group regardless of the nature or duration of the dietary regimen, time of blood collection, or subsequent tumor-bearing status. These data suggest that decreased serum insulin-like growth factor I/somatomedin C and insulin levels with CR and their complex interactions in vivo may play a role in the inhibition of mammary tumor promotion by CR.

Simultaneous isolation of insulin-like growth factors I and II from adult sheep serum.

Ovine insulin-like growth factors I and II (oIGF-I and oIGF-II) have been purified from adult sheep serum. oIGF-II-like receptor-binding activity and IGF-I-like immunoactivity were enriched on SP-Sephadex C-25, then purified using HPLC in the presence of a variety of counter ions. IGF-I- and IGF-II-like activities were separated using HPLC in the presence of 0.2% tetrabutylammonium phosphate at pH 7.0. The final recovery of oIGF-I was 82.6 micrograms from 3.2 litres of adult sheep serum (a yield of 17.6%), and the recovery of oIGF-II was 388 micrograms (a yield of 13.3%). Both IGF preparations were considered to be homogeneous as judged by single sharp peaks during analytical HPLC, and unique N-terminal amino acid sequences. Purified ovine IGFs had molecular weights similar to that of other IGFs (approximately 7000), and the first 30 N-terminal amino acids of both peptides were identical to their human counterparts. The isoelectric points of oIGF-I (pI approximately 8.2) and oIGF-II (pI approximately 6.8) were similar to those of human (h) IGFs (hIGF-I pI approximately 8.2; hIGF-II pI approximately 6.5), and the overall amino acid content of the ovine IGFs was also similar to that of IGFs from other species. oIGF-II preparations from fetal sheep and from adult sheep appeared to be identical. The isolation procedure represents one of general utility that can be easily modified to facilitate the isolation of recombinant IGFs from culture fluid.

Effect of rapid normalization of plasma glucose levels on microvascular dysfunction and polyol metabolism in diabetic rats.

Effects of rapid normalization of plasma glucose levels (by insulin infused via Alzet pumps implanted intraperitoneally) on plasma insulin-like growth factor I (IGF-I) levels, granulation tissue polyol levels, and vascular permeation by 125I-labeled albumin were examined in male Sprague-Dawley rats with streptozocin-induced (60-65 mg/kg) diabetes. Two days after implantation of pumps, plasma insulin levels were twice normal levels and remained elevated (1.4-2.5 times normal) throughout the remainder of the study. Plasma glucose levels and granulation tissue polyol levels were normalized within 2 days after initiation of insulin treatment. Plasma IGF-I levels were significantly increased (2 times) by 2 days, but were not normalized until 7 days. In contrast, 125I-albumin permeation normalized at a much slower relatively linear rate and was still not completely normal after 14 days of insulin treatment. In view of 1) previous studies demonstrating that diabetes-induced increases in 125I-albumin permeation in this tissue are linked to increased metabolism of glucose to sorbitol and 2) the rapid normalization of tissue polyol levels in this study, the relatively linear rate of normalization of vascular permeability over 14 days in these studies suggests that impaired vascular barrier functional integrity in this model is mediated by structural and/or functional vascular alterations associated with sustained increased polyol metabolism rather than by increased polyol levels per se and/or by readily reversible functional and metabolic alterations associated with acute increases in polyol metabolism.(ABSTRACT TRUNCATED AT 250 WORDS)

Antibodies to insulin-like growth factor I receptors in diabetes and other disorders.

A newly developed immunoprecipitation assay, with 125I-labeled highly purified human placental insulin-like growth factor I (IGF-I) receptor, was used to search for IGF-I-receptor antibodies in human sera. Eleven of 141 patient sera tested (7.8%) immunoprecipitated labeled IGF-I receptor. Immunoprecipitation was comparable with sera and IgG prepared from these sera. Seven of the 11 sera (3 of 31 with rheumatic disorders, 3 of 48 with non-insulin-dependent diabetes, and 1 of 52 with insulin-dependent diabetes) failed to inhibit IGF-I binding to human placental membranes and thus contained non-binding-inhibitory IGF-I-receptor antibodies. Their pathophysiological function remained uncertain. The remaining 4 sera (2 of 3 with type B severe insulin resistance, 1 of 7 with polycystic ovary syndrome (PCO), and 1 of 31 with rheumatic disorders) inhibited IGF-I binding. Plasma IGF-I concentrations were elevated (663 and 802 ng/ml, respectively) in 2 patients (1 with PCO and another with systemic lupus erythematosus) with binding-inhibitory IGF-I-receptor antibodies, suggesting IGF-I resistance that was probably mediated by the IGF-I-receptor antibodies. In conclusion, we identified two species of human IGF-I-receptor antibodies. Sera from 7 of 141 patients tested contained IgG autoantibodies that bound to the IGF-I receptor at a locus different from the IGF-I binding site and did not inhibit IGF-I binding. Sera from 4 of 141 patients contained antibodies that bound to the IGF-I receptor at or near the IGF-I binding site, inhibited IGF-I binding, and probably caused IGF-I resistance.

Degradation of insulin and insulin-like growth factors by enzyme purified from human erythrocytes. Comparison of degradation products observed with A14- and B26-[125I]monoiodoinsulin.

An insulin-degrading enzyme has been purified from human erythrocytes. This enzyme degraded 125I-labeled insulin-like growth factor I (IGF-I) more slowly than 125I-IGF-II and degraded IGF-II more slowly than 125I-insulin. The time course of 125I-insulin degradation suggested the presence of intermediates, each of which was itself shown to be a substrate for the enzyme. One of these intermediates appeared to be made up entirely of B-chain residues and had HisB10 as its NH2-terminal. The final major radiolabeled degradation product of A14-[125I]monoiodoinsulin was a peptide with TyrA14 at the A-chain NH2 terminal. This peptide could be reduced with dithiothreitol, suggesting that it contained amino acid residues from both A- and B-chains. It was partially precipitated by trichloroacetic acid and anti-insulin antibody but bound poorly to IM-9 lymphocytes. The final major degradation product of B26-[125I]monoiodoinsulin was a peptide whose NH2-terminal was TyrB26 and could not be reduced by dithiothreitol. It was partially precipitated by anti-insulin antibody but was precipitated poorly, if at all, by trichloroacetic acid and bound poorly to IM-9 lymphocytes. The results show that this enzyme degraded insulin by sequential cleavage of peptide bonds on both A- and B-chains. We identified LeuA13-TyrA14, SerB9-HisB10, and PheB25-TyrB26 as three of the bonds that are cleaved.

Nutrition and somatomedin. XVII. Circulating somatomedin C during treatment of diabetic ketoacidosis.

Although somatomedin levels may be low in animals with experimental diabetes, values in humans have generally been normal. Because humans with severely decompensated diabetes have not been studied, we characterized somatomedin C responses during metabolic restoration in 21 patients with diabetic ketoacidosis. Somatomedin C values were compared with those of 25 outpatient controls. Somatomedin C in controls (mean +/- SE) was 0.72 +/- 0.09 U/ml, 69% of the mean sex-adjusted normal level; 28% of patients had values below the lower limit of normal. In ketoacidotic subjects, initial somatomedin C was 0.43 +/- .06 U/ml, 33% of the mean normal level; 52% of subjects had somatomedin C below the lower limit of normal. Initial levels in ketoacidotic subjects were unrelated to presenting levels of glucose, bicarbonate, ketones, or blood urea nitrogen but were significantly lower in patients of less than ideal body weight (0.30 vs. 0.58 U/ml, P less than .03). Presenting levels of somatomedin C in ketoacidotic subjects were significantly lower than in controls (P less than .02). During insulin-infusion therapy, somatomedin C rose to a peak of 1.16 +/- 0.21 U/ml after 28 h, significantly higher than initial levels (P less than .05). With continued subcutaneous insulin, somatomedin C fell to a nadir after an additional 22 h then rose more slowly to a final value of 0.75 +/- 0.12 U/ml, significantly higher than the nadir (P less than .05) but lower than peak values; final values in the ketoacidotic subjects were comparable to outpatient somatomedin C levels.(ABSTRACT TRUNCATED AT 250 WORDS)

Insulin-like growth factors in vitreous. Studies in control and diabetic subjects with neovascularization.

Vitreous and serum were obtained at the time of vitrectomy from 23 diabetic subjects with proliferative retinopathy and from 8 nondiabetic subjects. The mean concentration of IGF-I in vitreous from diabetic patients with neovascularization was 6.3 +/- 0.93 versus 2.7 +/- 0.96 ng/ml. Chi-square and rank analysis indicated that higher concentrations of IGF-I occurred in diabetic vitreous (P less than 0.01 by both analyses). IGF-II concentrations in vitreous of control and diabetic subjects were not significantly different. A positive correlation existed between the concentrations of IGF-I and IGF-II in vitreous and their concentrations in serum in diabetic subjects, but not in control subjects. When vitreous concentrations of IGF-I were calculated for diabetic subjects studied previously with rapid acceleration of retinal disease, these concentrations varied from 20 to 30 ng/ml. The concentrations of IGF-I in the vitreous of most diabetic subjects with severe neovascularization are thus in the range known to stimulate cellular differentiation and growth in several systems. Whether they do so in the eye, and thus contribute to the development of retinopathy, remains to be determined.

Sex steroid dependency of diabetes-induced changes in polyol metabolism, vascular permeability, and collagen cross-linking.

The effects of castration on diabetes-induced increases in collagen cross-linking and vascular permeability and on polyol levels in new granulation tissue formed after induction of streptozocin (STZ) diabetes were examined in male Sprague-Dawley rats. New granulation tissue formation was induced by implanting sterile polyester fabric subcutaneously (s.c.) at the time of STZ injection 3 wk before assessment of vascular permeability and collagen cross-linking. Castration was performed 10 days before implanting the fabric. The characteristic increases in collagen cross-linking (manifested by decreased solubility in 0.5 M acetic acid) and in albumin permeation of the vasculature seen in intact diabetic rats were completely prevented by castration. Net collagen accumulation was not affected by diabetes or castration. Castration also markedly diminished diabetes-induced increases in tissue levels of sorbitol and completely prevented the decreases in tissue levels of myo-inositol and scyllo-inositol observed in intact diabetic rats, but had no effect on serum glucose levels, nonenzymatic glycosylation of plasma and granulation tissue proteins, or plasma somatomedin-C levels. The demonstration that castration prevents diabetes-induced increases in vascular permeability and collagen cross-linking as well as imbalances in tissue levels of sorbitol, myo-inositol, and scyllo-inositol in this model indicates that all of these changes are sex steroid-dependent phenomena. While the pathogenesis of these vascular permeability and collagen cross-linking changes is clearly multifactorial, these new findings: indicate that the role of sex steroids in the development of late complications of diabetes may be far more important than hitherto suspected, and suggest an explanation for the clinical observation that diabetic complications are uncommon in prepubertal diabetic subjects regardless of duration of diabetes.

Relationships between growth factors (somatomedin-C and growth hormone) and body development, metabolic control, and retinal changes in children and adolescents with IDDM.

We used the radioimmunoassay (RIA) method to determine somatomedin-C (SmC) basal values in 59 diabetic children and adolescents (20 prepubertal and 39 pubertal subjects; age range 2.75-20.16 yr; duration of diabetes 0.08-15.83 yr) and in 274 control subjects. In comparing diabetic subjects with controls, we considered only those 50 diabetic subjects who were age matched with the controls, i.e., those not over 16 yr chronological age. SmC basal levels in pubertal diabetic patients were no different from those of pubertal age-matched control children, whereas in prepubertal diabetic patients SmC was significantly lower than in the respective control children (P less than .001). No correlation was found between the z score for SmC (i.e., the number of standard deviations each SmC level is from the age- and sex-normalized mean) and duration of disease, velocity standard deviation score, severity of fluoroangiographic retinal changes, basal C-peptide values and HbA1 levels. No differences were encountered in mean SmC and SmC z-score values in the separate groups of poorly, fairly, and well-controlled diabetic children, in the groups with and without residual pancreatic activity, and in the group with and without retinal changes. In 16 of the pubertal diabetics and in 15 pubertal controls, serum glucose, growth hormone (GH), and SmC concentrations were determined during the night. The integrated nocturnal secretion of SmC was no different in diabetics than in controls, whereas the integrated nocturnal secretion of GH was significantly (P less than .025) higher in diabetics than in controls.(ABSTRACT TRUNCATED AT 250 WORDS)

Coordinate decrease of tissue insulinlike growth factor I posttranscriptional alternative mRNA transcripts in diabetes mellitus.

In these studies, we examined the effect of excess levels of growth hormone (GH) on rat insulinlike growth factor I (IGF-I) gene expression in streptozocin-induced diabetes mellitus. A solution hybridization/RNase protection assay was used to simultaneously quantitate the relative tissue content of the variant IGF-I mRNA species arising from alternative splicing in the region encoding the COOH-terminal extension E-peptide (IGF-Ia and IGF-Ib). IGF-Ia and IGF-Ib mRNAs were markedly decreased in liver, kidney, and lung tissues of diabetic rats. Although GF stimulates IGF-I gene expression, chronic GH excess from implanted somatomammotropic tumors did not appropriately induce tissue IGF-I mRNA content in diabetic animals. Treatment of diabetic rats with insulin for 1 wk restored basal and GH-stimulated IGF-Ia and IGF-Ib mRNA content toward that present in tissues of nondiabetic rats. The ratio of IGF-Ia to IGF-Ib mRNA remained relatively constant for each tissue and was not affected by the diabetic state, chronic GH hyperstimulation, or insulin therapy, suggesting that posttranscriptional splicing is not a regulated event in these conditions. Thus, both circulating IGF-I levels and tissue IGF-I gene expression are profoundly decreased in this model of experimental diabetes. Diminished tissue availability of IGF-I from endocrine and/or paracrine sources may be responsible for the growth retardation seen in uncontrolled diabetes mellitus.

Nutrition and somatomedin. XIX. Molecular regulation of insulin-like growth factor-1 in streptozotocin-diabetic rats.

Poor growth in diabetes involves low circulating levels of somatomedins/insulin-like growth factors (IGFs), largely reflecting decreased growth factor release by the liver. To define regulatory mechanisms, circulating IGF-1 was compared with levels of a high mol wt putative hepatic IGF-1 precursor and hepatic IGF-1 mRNA in a model of progressive severity of diabetes in rats. Streptozotocin administered at 36, 72, 144, and 288 mg/kg produced graded metabolic decompensation 2 days later, from minimal hyperglycemia with continued weight gain at 36 mg/kg, to marked hyperglycemia, ketonemia, and weight loss at 288 mg/kg (all P less than 0.001). Total serum IGF-1 measured by RIA was unchanged with the 36 and 72 mg/kg doses of streptozotocin (471 +/- 19 and 439 +/- 27 ng/ml, respectively, vs. 517 +/- 27 ng/ml in controls) despite serum glucose greater than 400 mg/dl. With streptozotocin 144 and 288 mg/kg, serum IGF-1 fell to 131 +/- 27 and 142 +/- 10 ng/ml, respectively (both P less than 0.005 vs. controls). Serum IGF-1 was correlated strongly with serum beta-hydroxybutyrate and body weight (r = -0.88 and 0.91, respectively, P less than 0.0001), and less strongly with serum glucose (r = -0.59, P less than 0.0002). Extractable hepatic content of a high mol wt form of immunoreactive IGF-1 (a putative precursor) was unchanged at the two lowest doses of streptozotocin (68 +/- 4 and 83 +/- 9 ngeq/g vs. 67 +/- 4 in controls), but decreased to 16 +/- 3 and 29 +/- 4 ng/g at the two highest doses (both P less than 0.001 vs. controls).(ABSTRACT TRUNCATED AT 250 WORDS)

The effects of muscle-building exercise on vitamin D and mineral metabolism.

Exercise and muscle strength are important determinants of bone mass. Studies were carried out in normal young adult white males to determine the effects of exercise on vitamin D and mineral metabolism. Fourteen men who had engaged in regular muscle-building exercises for at least 1 year and 14 age-matched controls (age range, 19-36 year) were hospitalized on a metabolic ward and were given a constant daily diet estimated to contain 400 mg of calcium, 900 mg of phosphorus, 110 mEq of sodium, 65 mEq of potassium, and 18 mEq of magnesium. Body weight averaged 78 +/- 3 kg in the exercisers and 72 +/- 2 kg in the controls (NS). Serum calcium, ionized calcium, phosphate, magnesium, somatomedin-C, and immunoreactive parathyroid hormone (PTH) were not different in the two groups, whereas serum Gla-protein (39 +/- 5 vs. 24 +/- 2 ng/ml, p less than 0.01), 25-hydroxyvitamin D (23 +/- 2 vs. 16 +/- 2, p less than 0.05) and 1,25-dihydroxyvitamin D [1,25(OH)2D] (40 +/- 2 vs. 29 +/- 2 pg/ml, p less than 0.01) were higher in the exercisers than in the controls. Urinary calcium, phosphorus, sodium, potassium, creatinine clearance, and norepinephrine were not different in the two groups, whereas urinary magnesium (12.6 +/- 1.0 vs. 9.4 +/- 0.5 mEq/d, p less than 0.01) and urinary cyclic adenosine 3',5'-monophosphate (cyclic AMP) (2.52 +/- 0.19 vs. 1.72 +/- 0.20 nM/dl glomerular filtrate, p less than 0.01) were higher in the exercisers than in the controls.(ABSTRACT TRUNCATED AT 250 WORDS)

Elevated plasma IGF 1 levels in stags prevented from growing antlers.

We have previously shown that plasma IGF 1 concentration is positively correlated with the rate of antler growth and have proposed that IGF 1 is stimulatory for antler growth in the red deer stag. Therefore to partly resolve the question of whether the IGF 1 was of circulating or local origin in relation to its effect on antler growth, we surgically prevented stags from growing antlers. We recorded significantly elevated plasma levels of IGF 1 in the non-antlered stags compared with normal antlered stags during the antler growth periods. This result is consistent with a hypothesis that the antler is a target organ for IGF 1 and that prevention of antler growth removed a population of IGF 1 receptors. IGF 1 is already known to stimulate body growth but this work points strongly to the possibility that plasma IGF 1 may stimulate individual organ growth in an endocrine manner.

An improved method for the purification of human insulin-like growth factors I and II.

Insulin-like growth factors (IGFs) I and II have been purified from Cohn fraction IV-1 of human plasma. After acid-ethanol extraction, the consecutive use of conventional gel filtration and reverse phase liquid chromatography has permitted the rapid isolation of these polypeptides. Purification was monitored by the use of specific RIAs. In both chromatography systems, separation was optimized by performing it on the same stationary phase but successively with mobile phases of different pH or different solute selectivity. The two polypeptides were shown to be pure by their unique amino acid composition, particularly by the absence of specific amino acids (histidine, tryptophan, and methionine), and their unique amino-terminal sequences. In addition, the lack of cross-contamination of the two growth factors with each other was established by the unique isoelectric focusing patterns of IGF-I at pI 8.25 and IGF-II at pI 6.5. From 900 g Cohn fraction IV-1, which is equivalent to 66 liters human plasma, approximately 100 micrograms of each IGF can be obtained by our procedure, which can easily be carried out in a clinical research laboratory.

Insulin-like growth factors I and II in maternal and fetal guinea pig serum.

The role of insulin-like growth factors (IGFs) in fetal development has been the subject of much speculation. We undertook studies of maternal and fetal IGF I and II in the guinea pig because the long gestation period and greater size of the fetuses permitted blood sampling over a longer period of gestation and maturation than is possible in the rat. Acid gel filtrates of fetal and maternal serum were prepared, and the IGF I was measured by RIA; IGF II was measured by rat placental membrane radioreceptor assay. Fetal IGF I levels were lower than maternal levels from the 33rd day of estimated gestation to term. Fetal IGF II levels from the 33rd day to the 49th day of gestation were not significantly different from those of maternal serum [1597 +/- 377 (SE) ng/ml vs. 1295 +/- 224] ng/ml. Very high levels of IGF II, in excess of 5000 ng/ml, were observed in fetuses at 50, 55, and 60 days of gestation. Thereafter, fetal IGF II levels fell markedly before term. Fetal and maternal IGFs after 49, 50, 60, and 65 days of pregnancy were compared by isoelectric focusing. The guinea pig normally has two major basic peaks of IGF I, which were present both in maternal and fetal serum. Most maternal and fetal guinea pig sera contained only a single, slightly acidic peak of IGF II. No evidence of a unique fetal IGF was detected by our methods. The very high levels of IGF II reached in fetal guinea pig sera suggest that it may have a role in fetal development.

Genetic selection for insulin-like growth factor-1 in growing mice is associated with altered growth.

Substantial responses in the 6-week and mature body-weights of mice occurred after 7 generations of selection for or against plasma levels of Insulin-like Growth Factor-1 (IGF-1). Plasma levels of IGF-1 were also significantly different after 7 generations of selection (high line = 85 +/- 2 ng/ml, low line = 58 +/- 2 ng/ml). The average 6-week weight in the line selected for high plasma IGF-1 was 22.5 +/- .2 g compared with 18.5 +/- .2 g in the low plasma IGF-1 line, after 7 generations of selection. The difference between lines was maintained at 20 weeks of age. These data provide further evidence for the roles of IGF-1 in the regulation of somatic growth and as a mediator of a genetic component of growth.

The ontogeny and regulation of a 31,000 molecular weight insulin-like growth factor-binding protein in fetal porcine plasma and sera.

Studies were conducted to determine the presence, quantity, and regulation of insulin-like growth factor (IGF)-binding proteins in porcine plasma and sera. The size distribution of IGF-binding protein was determined by affinity cross-linking to [125I]IGF-I, and the binding activity was quantified by a polyethylene glycol precipitation procedure. The major [125I]IGF-I-binding protein complex was of 38,000 mol wt (Mr) in all porcine sera or plasma tested. Binding to this protein was inhibited by excess unlabeled IGF-I, but not by insulin. When plasma samples from fetuses at 45, 70, 90, and 110 days gestation were cross-linked to [125I]IGF-I, there was an increase in the concentration of the 38,000 Mr complex with advancing gestational age, whereas plasma from the mothers of the age-matched fetuses showed little change in the amount of the 38,000 Mr complex. When the binding activity of the fetal plasma was quantified, there was a significant increase in binding activity between 45 and 110 days gestation from 30.5 +/- 3.3% to 77.5 +/- 3.6% (+/- SE) of the assay maximum. This increase was not due to a decrease in binding protein saturation, since the endogenous IGF-I levels (quantified after acid gel filtration chromatography) also increased with advancing gestational age, from 176 to 458 mU/ml. Blood samples were collected from porcine fetuses at 110 days gestation that had either been decapitated or spinal cauterized at 45 days gestation. Decapitation decreased the amount of the 38,000 Mr complex that could be detected by affinity labeling. In contrast, spinal cauterization had no effect on the amount of the 38,000 Mr complex in fetal plasma compared to that in control fetal plasma. These results were verified in the polyethylene glycol assay. These studies show that the amount of the 38,000 Mr IGF-binding protein complex in pig plasma is developmentally regulated and suggest that a pituitary or neural factor may be an important variable in the control of its plasma concentrations.