Biomedical subjects

D R Clemmons

Publications and source records attributed to D R Clemmons.

At least 163 records · Page 9Linked to original sources

PubMed1991

Evaluation of the developmental and nutritional changes in porcine insulin-like growth factor-binding protein-1 and -2 serum levels by immunoassay.

Porcine serum contains five insulin-like growth factor-binding proteins (IGFBPs), whose regulation has been studied by ligand blotting. To more accurately quantify changes in two specific forms of IGFBP a heterologous RIA for porcine (p) IGFBP-2 was developed, and IGFBP-1 levels were analyzed by immunoblotting. By RIA, postnatal hypophysectomy caused a 7-fold increase in serum pIGFBP-2 levels compared to controls (2,622 +/- 378 vs. 382 +/- 10 ng/ml, respectively). Fetal pIGFBP-2 levels were higher at 110 vs. 45 days gestation (1,074 +/- 214 vs. 418 +/- 30 ng/ml, respectively), rose to 1,905 +/- 167 ng/ml within 12 h after birth, then decreased to 1,010 +/- 10 ng/ml at 48 h. By immunoblot analysis, bovine IGFBP-2 antiserum reacted with a 34,000 mol wt (Mr) IGFBP and did not react with other forms of IGFBP detected by ligand blotting. Serum levels of the 34,000 Mr IGFBP, as detected by ligand blot analysis, are decreased when neonatal pigs are fasted for 48 h. In contrast, by RIA, pIGFBP-2 concentrations increased 4-fold. Immunoblots of these sera showed two lower Mr (22,000 and 14,000 Mr) bands that did not bind either [125I]IGF-I or [125I]IGF-II and were distinct from a smaller (20,000 Mr) IGFBP which bound only [125I]IGF-II. These two bands were increased in serum of 48-h fasted compared to fed piglets, suggesting that they are proteolytic fragments of pIGFBP-2. In vitro incubation of 48-h fasted pig serum with intact IGFBP-2 failed to reveal proteolytic fragments, indicating that the IGFBP-2 fragments were not generated by a protease that was released into the serum. Analysis performed with human IGFBP-1 antiserum revealed a 29,000 Mr immunoreactive band whose abundance was increased by either postnatal hypophysectomy or fasting. No fragments of IGFBP-1 were found in any serum tested. We conclude that heterologous antibodies can be used to identify and quantify IGFBP-1 and IGFBP-2 in porcine serum. Changes in pIGFBP-2 levels measured during fasting are due to the combination of changes in intact 34,000 Mr IGFBP-2 and smaller non-IGF-binding fragments. Changes in levels of specific forms of IGFBP as well as the presence of fragments have the potential to modulate the transport of IGF-I and -II out of the vasculature.

Animal Nutritional Physiological Phenomena↗

PubMed1991

Variables controlling the secretion of insulin-like growth factor binding protein-2 in normal human subjects.

Insulin-like growth factor binding protein-2 (IGFBP-2) is one of a family of IGFBPs that are present in extracellular fluids, and binds both IGF-II and IGF-I with high affinity. These studies were conducted to determine the nutritional and hormonal variables that regulate plasma IGFBP-2 concentrations in humans. The mean plasma IGFBP-2 concentration for 38 normal adult subjects was 150 +/- 61 micrograms/L and was 4.7-fold greater than their mean fasting IGFBP-1 value. Mean IGFBP-2 values in cord sera of 26 normal term infants was 3.8-fold greater than the normal adult mean value. Likewise, the mean value for 44 hypopituitary adults was increased 2-fold compared to normal. There was no suppression of IGFBP-2 values in acromegaly. Normal adult subjects showed minimal fluctuations (less than 2-fold changes) in plasma IGFBP-2 concentrations during a 48-h sampling period. These changes were significantly less than the changes that occurred in plasma IGFBP-1 during the same interval. Plasma IGFBP-2 did not change significantly post prandially or after a glucose infusion. Extreme insulin deficiency, after 9 days of fasting, was associated with a 1.7-fold increase in plasma IGFBP-2. Administration of GH, which is known to cause a major decrease in plasma IGFBP-1 and in IGFBP-2 in hypophysectomized animals, did not result in a change in calorically restricted normal adult subjects, suggesting that a normal caloric intake is required for GH to suppress IGFBP-2. In summary, these results show that plasma IGFBP-2 is regulated differently than IGFBP-1. Acute stimulation of insulin secretion does not suppress IGFBP-2, and there is much less daily fluctuation compared to IGFBP-1. These findings suggest that plasma IGFBP-2 levels are more stable than IGFBP-1, and therefore IGFBP-2 may serve as a larger reservior that is available for IGF transport.

Acromegaly↗

PubMed1991

The effect of growth hormone on weight gain and pulmonary function in patients with chronic obstructive lung disease.

Thirty to 60 percent of patients with chronic obstructive pulmonary disease (COPD) are malnourished, and this affects ventilatory muscle function and prognosis for survival adversely. We studied the effect of growth hormone (GH) in malnourished patients with COPD (n = 7, mean FEV1 of 1.1 +/- 0.2 L, 45 +/- 7 percent of predicted values; less than 90 percent of ideal body weight). The subjects received a balanced diet of 35 kcal/kg with 1 g of protein/kg for 1 week (pre-GH). During the following three weeks, they received the same diet plus subcutaneous injections of recombinant methionyl human GH (0.05 mg/kg daily). The subjects had no significant weight gain during the week of diet alone (0.07 +/- 0.11 kg, p = NS) but they had substantial weight gain during the first week of GH treatment (1.37 +/- 0.23 kg, p less than 0.01). Nitrogen balance also improved during GH treatment (+1.6 +/- 0.7 g/day on diet alone vs +3.8 +/- 0.5 g/day on diet plus GH, p less than 0.02). Maximal inspiratory pressure improved by 27 +/- 8 percent after GH treatment (p less than 0.02). No significant adverse effects occurred. Further study of the potential usefulness of GH in COPD utilizing a placebo-controlled trial is warranted.

Aged↗

PubMed1991

Role of peptide growth factors in development of macrovascular complications of diabetes.

Peptide growth factors provide an important means of coordinating the growth of cells within tissues and organs. Although their role in controlling cell growth is not well understood, they have been implicated in derangements of cellular proliferation that occur in diabetes, e.g., mesangial cell hyperplasia and atherosclerosis. Because several growth factors have been structurally characterized and the cell types on which they act identified, research is focusing on developing model systems to determine whether they are involved in the pathogenesis of specific disease states. New techniques, i.e., in situ hybridization, gene transfection, and detailed structural analysis of proteins, have made it possible to define both changes in the relative abundance of specific growth factors and potential changes in their actions in specific disease states. These techniques are being applied in diabetes research and will make it possible to determine the alterations that have occurred in growth factor synthesis and growth factor-matrix protein interaction and cell-type-specific alterations in cell growth that occur after loss of normal glucose homeostasis. The findings from these types of analyses should lead to a better understanding of how the complications of diabetes develop and rational strategies to control their effects.

Diabetes Mellitus↗

PubMed1991

Insulin-like growth factor binding proteins: roles in regulating IGF physiology.

Insulin-like growth factor binding proteins (IGFBPs) are soluble proteins present in in extracellular fluids. They have high affinity for IGF-I and -II. Blood concentrations are controlled by nutrition and by hormones in a manner that in most, but not all, instances correlates with plasma concentrations of IGF-I or -II. IGF binding proteins are secreted by a range of cell types in a manner that may serve to modulate the functions of the growth factors in a pericellular environment. IGF binding proteins cxan modify IGF interaction with the type I receptor and may thereby alter IGF signal transduction through this transmembrane signalling unit. Binding proteins may also act as inhibitors or potentiators of biological responsiveness and thereby directly cell type specific responses.

Carrier Proteins↗

PubMed1990

Regulation of breast cancer growth by insulin-like growth factors.

The IGFs may be important autocrine, paracrine or endocrine growth factors for human breast cancer. IGF-I and II stimulate growth of cultured human breast cancer cells. IGF-I is slightly more potent, paralleling its higher affinity for the IGF-I receptor. Antibody blockade of the IGF-I receptor inhibits growth stimulation induced by both IGFs, suggesting that this receptor mediates the growth effects of both peptides. However, IGF-I receptor blockade does not inhibit estrogen (E2)-induced growth suggesting that secreted IGFs are not the major mediators of E2 action. Several breast cancer cell lines express IGF-II mRNA by both Northern analysis and RNase protection assay. IGF-II activity is found in conditioned medium by radioimmuno and radioreceptor assay, after removal of somatomedin binding proteins (BP) which are secreted in abundance. IGF-I is undetectable. BPs of congruent to 25 and 40 K predominate in ER-negative cell lines while BPs of 36 K predominate in ER-positive cells. Blockade of the IGF-I receptor inhibits anchorage-independent and monolayer growth in serum of a panel of breast cancer cell lines. Growth of one line (MDA-231) was also inhibited in vivo by receptor antibody treatment of nude mice. The antibody had no effect on growth of MCF-7 tumors. These data suggest that IGFs are important regulators of breast cancer cell proliferation and that antagonism of this pathway may offer a new treatment strategy.

Antineoplastic Agents↗

PubMed1990

Discrete alterations of the insulin-like growth factor I molecule which alter its affinity for insulin-like growth factor-binding proteins result in changes in bioactivity.

Insulin-like growth factor (IGF)-binding proteins (BPs) bind IGF-I and IGF-II with high affinity. They are present in extracellular fluids and modulate the interactions of their ligands with the type 1 IGF cell surface receptor. These studies utilized IGF-I analogs that have reduced binding affinity for either the type 1 IGF receptor or binding proteins to study the ligand specificity of IGF-BP-1 and the role of IGF-BP-1 in modulating the biological activity of IGF-I. The data indicate that the regions of IGF-I which are responsible for binding to IGF-BP-1 and to human serum-binding proteins are distinct but overlapping and are clearly distinct from the type I receptor binding sites. In the absence of exogenously added IGF-BP-1, the analogs with reduced affinity for IGF-BP-1 are more potent than IGF-I in stimulating DNA synthesis by porcine aortic smooth muscle cells. In contrast, when cells are concomitantly exposed to IGF-BP-1, two of the analogs with reduced affinity for binding protein give only 40-65% of the maximal IGF-I response. [Leu24, 1-62]IGF-I, which has a 100-fold reduced affinity for the type 1 IGF receptor, gave a value that was 62% of the maximal IGF-BP-1 potentiated response. A second biological response, that of stimulating binding protein secretion by IGF-I, was also examined. [Leu24, 1-62]IGF-I is more potent than IGF-I whereas the activity of the analogs with lower affinity for IGF-BP-1 is significantly reduced. Thus, the ability to activate DNA synthesis and binding protein secretion maximally in the presence of IGF-BP-1 is dependent on the affinity of IGFs for both type 1 receptors and binding proteins.

Amino Acid Sequence↗

PubMed1990

Insulin-like growth factor (IGF) binding to human fibroblast and glioblastoma cells: the modulating effect of cell released IGF binding proteins (IGFBPs).

The cell surface of human fibroblasts contains not only type I IGF receptors but at least two forms of IGFBPs. Studies were undertaken to analyze the mechanisms by which these IGFBPs alter IGF-I-cell surface interactions. Human fetal fibroblasts (GM10) and a human glioblastoma cell line (1690) were chosen for analysis. During assays to quantify [125I]-IGF-I binding, both cell lines were shown to release IGFBPs into the binding assay buffer. Under equilibrium conditions, [125I]-IGF-I preferentially associates with IGFBPs in the assay buffer (up to 40% of the [125I]-IGF-I added) since they have a higher affinity than type I IGF receptors or IGFBPs associated with the cell surface. Likewise the addition of increasing concentrations of unlabeled IGF-I results in preferential competition for binding to assay buffer IGFBPs. This results in a repartitioning of the [125I]-IGF-I that is bound to assay buffer IGFBPs onto cell surface binding sites. The degree of repartitioning is quantitatively related to the amount of [125I]-IGF-I bound to released IGFBPs. When cultures are exposed to cycloheximide before the binding assay, both the amount of IGFBPs that are released into the assay buffer and the amount of [125I]-IGF-I that is repartitioned are decreased. In contrast when [Gln3, Ala4, Tyr15, Leu16]-IGF-I ([QAYL]-IGF-I, an IGF analog that has unaltered affinity for type I IGF receptors) is iodinated and tested, the competition curve with unlabeled IGF-I shows no repartitioning effect. This form of IGF can be used to quantify type I receptor number independent of the presence of IGFBPs. IGF-I and the [QAYL]-IGF-I compete equally with the [125I]-[QAYL]-IGF-I for binding to cell surfaces, whereas unlabeled [QAYL]-IGF-I is greater than 25-fold less potent compared to IGF-I in competing with [125I]-IGF-I for cell surface binding. Specific binding of [125I]-[QAYL]-IGF-I to GM10 and 1690 cell surfaces is less than 20% of [125I]-IGF-I binding. These findings suggest that IGFBPs that are present on human fibroblast surfaces represent a large portion of the IGF binding sites. We conclude that the amount of IGFBPs released into assay buffer is a major determinant of the repartitioning of [125I]-IGF-I to cell surface binding sites and that both cell surface and assay buffer IGFBPs modulate type I IGF receptor binding.

Binding, Competitive↗

PubMed1990

A factor contained in plasma is required for IGF binding protein-1 to potentiate the effect of IGF-I on smooth muscle cell DNA synthesis.

One of the forms of the insulin-like growth factor (IGF) binding proteins present in human amniotic fluid has been shown to potentiate the growth-promoting effect of IGF-I markedly. This study was undertaken to determine the cellular and hormonal factors that modulate this potentiation and to determine whether this protein would potentiate the effects of other mitogens. Although the combination of the IGFBP-1 (20 ng/ml) and IGF-I (10 ng/ml) induced a five- to sixfold increase in DNA synthesis compared with IGF-I alone, this response required the simultaneous addition of IGF-I with 0.1% platelet-poor plasma (PPP). If PPP was omitted from the incubation medium, no increase above the effect that was obtained with IGF-I alone was noted. Substitution of cerebrospinal fluid (CSF) for PPP permitted a full mitogenic response, although substitution with amniotic fluid resulted in no enhancement. The factor contained in PPP was heat and acid stable. If the binding protein was co-incubated with fibroblast growth factor (FGF), platelet-derived growth factor (PDGF), or epidermal growth factor (EGF), a slight inhibition of the cellular response to each of these factors was detected. Co-incubation of IGF-I with the IGF-binding protein plus these other peptide growth factors resulted in no further enhancement of DNA synthesis above the level observed with IGF-I and the binding protein alone. Likewise, addition of plasma proteins such as transferrin or albumin did not result in a further enhancement of the DNA synthesis response to IGF-I plus binding protein, and these proteins could not substitute for PPP or IGFBP-1. Transient exposure of the cultures (2 hr) to the binding protein plus IGF-I resulted in a submaximal DNA synthesis response, and the binding protein had to be present continuously to achieve a maximal effect. These studies indicate that a factor contained in plasma and CSF is required for a maximal cellular response to IGFBP-1 plus IGF-I, and this factor does not appear to be a well-defined mitogen.

Animals↗

PubMed1990

Anabolic effects of growth hormone in obese diet-restricted subjects are dose dependent.

In previous studies growth hormone (GH) injections [0.1 mg/kg ideal body wt (IBW) every other day] produced significant increases in plasma insulin-like growth factor I (IGF-I) concentrations and nitrogen retention, which were attenuated when 12 kcal/kg IBW was ingested. The present study was done to determine whether doubling the GH dose would enhance its anabolic effects and facilitate fat loss. Eight women (33-83% over IBW) were fed 12 kcal/kg IBW for 14 wk. They received GH or vehicle injections, each for 5 wk during either weeks 2-6 or 9-13. GH improved nitrogen balance (GH, 3.6 +/- 123.2 mmol/d; vehicle, -132.0 +/- 117.9 mmol/d; means +/- SD; p less than 0.001). Plasma IGF-I increased from 32.1 +/- 9.6 to 79.4 +/- 22.1 nmol/L by day 5 of GH (p less than 0.001) and remained elevated until GH injections were discontinued. No significant effect of GH on mean body fat loss was observed. GH can induce significant anabolic responses even when caloric intake is decreased to 12 kcal/kg IBW. The degree and duration of these anabolic responses are dependent on the GH dose given.

Adolescent↗

PubMed1990

Differential regulation of insulin-like growth factor binding protein secretion from human decidual cells by IGF-I, insulin, and relaxin.

Several growth hormone-independent 25-31,000 kD insulin-like growth factor binding proteins (IGF-BPs) have been identified in plasma, extravascular fluids, and various cell-conditioned media. Cultured human decidual cells release three IGF-BPs with 24,000, 30,000, and 34,000 Mr. Using ligand blot analysis and an RIA for the 30,000-Mr form (IGF-BP-1), we examined the effects of IGF-I (10-1,000 ng/ml), insulin (10-10,000 ng/ml), and relaxin (10-250 ng/ml) on decidual cell IGF-BP release after 120 h of hormone exposure. IGF-I inhibited release of both IGF-BP-1 and the 24,000 Mr form. Inhibition of IGF-BP-1 release was noted after 48 h of treatment and was progressive throughout the subsequent 120 h. Insulin stimulated a fourfold increase in release of the 24,000-Mr protein while inhibiting IGF-BP-1 release comparable to IGF-I, alpha-IR3, a monoclonal antibody to the IGF-I receptor, blocked approximately 33% of the IGF-I response but had no effect on insulin-mediated IGF-BP-1 inhibition. Relaxin stimulated a 2.4-fold increase in release of the 24,000-Mr form and a 16-fold increase in the 30,000-Mr protein after 120 h. Stimulation of the 30,000-Mr protein was inhibited by the addition of cycloheximide (50 micrograms/ml). Both IGF-I and insulin also blocked the relaxin-mediated increase in IGF-BP-1. These studies suggest that three structurally related proteins differentially regulate IGF-BP secretion possibly via activation of distinct receptor subtypes.

Antibodies, Monoclonal↗

PubMed1990

Insulin differentially alters transcapillary movement of intravascular IGFBP-1, IGFBP-2 and endothelial cell IGF-binding proteins in the rat heart.

Insulin-like growth factor binding-proteins 1 and 2 (IGFBP-1, IGFBP-2) and endothelial cell IGF binding proteins (ECBP) were individually perfused through isolated beating rat hearts in the absence and presence of insulin. Insulin caused an increased movement of IGFBP-1 from the vascular space to tissues of the heart. Subendothelial content of IGFBP-1 was 110%, 126% (p less than .01) and 132% (p less than 0.05) of control hearts when perfused with 1, 10 and 100 ng/ml insulin, respectively. . In contrast, insulin treatment was associated with a decrease in ECBP content in cardiac tissue, being 83%, 62% (p less than 0.005) and 73% (p less than 0.05) of control when perfused with 1, 10 and 100 ng/ml insulin. The efflux of IGFBP-2 from the intravascular space was unaffected by insulin. The subendothelial tissue distribution of the transported binding proteins was not changed by insulin perfusion, with IGFBP-1 and IGFBP-2 localizing predominantly in cardiac muscle and ECBP having greater affinity for connective tissue elements. We conclude that in the perfused rat heart, insulin can differentially alter transcapillary movement of IGFBP-1, IGFBP-2 and endothelial cell IGF-binding proteins. Such insulin-facilitated changes could potentially mediate nutrient-dependent transport of IGF-I and IGF-II to peripheral tissues.

Animals↗

PubMed1990

Three distinct forms of insulin-like growth factor binding proteins are released by decidual cells in culture.

Recent studies have shown that human decidual explants synthesize a 25,272 dalton form of insulin-like growth factor binding protein (IGFBP-1) that is identical to the most abundant form of IGFBP detected in human amniotic fluid. To determine whether decidual cells also secrete other structurally distinct forms of IGFBP, conditioned medium obtained from human decidual cells was analyzed by ligand blotting and immunoblotting using antisera that were specific for IGFBP-1 or 2. IGFBP-2 is a form of binding protein that is structurally distinct from IGFBP-1. Ligand blotting analysis (which detects all forms of IGFBPs) showed that 3 forms of IGFBP with Mr estimates of 34,000, 30,000 and 24,000 were present in basal, unstimulated, conditioned media. Immunoblotting showed that the 30,000 Mr form reacted with an antibody that is specific for IGFBP-1, whereas the 34,000 Mr form reacted with an antibody that is specific for IGFBP-2. The 24,000 Mr band did not react with antisera to either IGFBP-1 or 2. Basal IGFBP-1 release from human decidual cultures, as measured by RIA, showed a significant decrease during the 5 day period from 34 +/- 1.6 on day 1 to 12 +/- 1.9 ng/ml on day 5. Exposure of the cells to (Bu)2cAMP (1.0 mM) for 5 days had no significant effect on IGFBP-1 release during the first day of exposure, but caused a progressive stimulation of release during the subsequent 4 days. By day 5, exposure to (Bu)2cAMP induced a 5- to 6-fold increase in IGFBP-1 release that was completely inhibited by exposure to cycloheximide. Exposure of the cells to cholera toxin (0.1 micrograms/ml) for 4 days caused a 3.7-fold increase in IGFBP-1 release. The stimulation of IGFBP-1 release by (Bu)2cAMP was completely inhibited by simultaneous exposure to insulin (100 ng/ml) and/or IGF-I (100 ng/ml) (day 5 control = 35 ng/ml, (Bu)2cAMP alone = 248 ng/ml, insulin + (Bu)2cAMP = 36 ng/ml, or IGF-I + (Bu)2cAMP = 10.1 ng/ml). Hydrocortisone and the phorbol ester (phorbol myristate acetate) caused no significant changes in basal IGFBP-1 release but prevented the decrease in the basal rate of release that occurred between day 1 and 5. Ligand blotting studies showed that the 24,000 Mr form of binding protein was also stimulated by (Bu)2cAMP (3.3-fold increase); however, cholera toxin and phorbol myristate acetate were without effect. In contrast, the release of the 34,000 Mr form was unaffected by exposure to any of these agents, suggesting that regulation of this form of IGFBP is distinct from the other 2 forms.(ABSTRACT TRUNCATED AT 400 WORDS)

Bucladesine↗

PubMed1990

Transcapillary permeability and subendothelial distribution of endothelial and amniotic fluid insulin-like growth factor binding proteins in the rat heart.

Insulin-like growth factor (IGF) binding proteins (IGFBP) were purified from conditioned media of cultured bovine endothelial cells (ECBP) and from human amniotic fluid (IGFBP-1), and then labeled by radioiodination. 125I-ECBP and 125I-IGFBP-1 were perfused through isolated beating rat hearts for 1 and 5 min, and the hearts fixed and analyzed for 125I-BP content and distribution. One to 4% of the perfused 125I-ECBP and 125I-IGFBP-1 crossed the capillary boundary. The ECBPs predominantly localized as intact 125I-BP in connective tissue elements of the heart with less 125I-BP in cardiac muscle. The ratio of 125I-ECBP in connective tissue: muscle (normalized to percent vol of these compartments) was greater than or equal to 10:1. In contrast, the IGFBP-1 had a greater affinity for cardiac muscle with ratios of 125I-IGFBP-1 in connective tissue:muscle of approximately 1:2. When 125I-IGF-I, in the absence of any BPs, was perfused through the hearts approximately 3-5% left the microcirculation and was found in subendothelial tissues. 125I-IGF-I localized primarily to cardiac muscle with a distribution of connective tissue:cardiac muscle of about 1:3. The findings in the isolated perfused heart were confirmed in intact animals. After 125I-IGFBP-1 was injected into anesthetized rats and allowed to circulate for 5 min, substantial radioactivity was associated with the heart. As in the isolated heart, the IGFBP-1 preferentially localized to cardiac muscle with a connective tissue:cardiac muscle ratio of 1:3. We conclude that IGFBPs produced by endothelial cells and the IGFBP-1 contained in amniotic fluid can cross the capillary boundaries of the rat heart, and that the ECBPs preferentially localize in connective tissue elements of the myocardium, whereas IGFBP-1 predominantly localizes in cardiac muscle.

Amniotic Fluid↗

PubMed1990

Regulation of serum insulin-like growth factor-I (IGF-I) and IGF binding proteins during rat pregnancy.

Serum concentrations of insulin-like growth factor-I (IGF-I) in rats are reduced dramatically in the latter half of pregnancy, decreasing from 1758 +/- 356 ng/ml at 12 days of pregnancy (mean +/- SD) to 761 +/- 192 ng/ml at 15 days. After parturition, IGF-I increases to nonpregnant values in 4 days. Using ligand blotting, we have demonstrated that most of the serum IGF binding proteins (IGFBPs) are concurrently reduced during pregnancy. IGFBP-3, the predominant IGFBP in nonpregnant serum, is reduced to 1.3% of nonpregnant values by 21 days of pregnancy and begins to rise within 1 h postpartum (PP). The sera of 21-day pregnant (but not nonpregnant) rats degrade IGFBP-3 in vitro, and this degradation is prevented by the protease inhibitor antipain. Decreased serum IGF-I concentrations during pregnancy, therefore, may result from reduced IGFBP-3 concentrations causing increased IGF-I clearance. In addition, steady state IGF-I mRNA and peptide levels in liver are decreased in 21-day pregnant rats (37% and 42% of 4 day PP levels, respectively), suggesting that decreased synthesis of IGF-I may also lead to lower serum IGF-I concentrations. After bolus injection, [125I]IGF-I is cleared from the serum of pregnant rats nearly 5 times faster than that of 4 day PP rats (1.21 vs. 0.25 ml/min/kg, respectively). Urinary clearance is relatively insignificant (less than 4%), and [125I]IGF-I does not cross the placenta. The intermediate distribution phase of IGF-I is slower in pregnant rats than in PP rats (t1/2 alpha, 17.1 vs. 5.4 min), whereas the terminal elimination of IGF-I is twice as fast (t1/2 beta, 228.1 vs. 106.4 min). The prolonged IGF-I distribution phase in the pregnant rats may result from decreased concentrations of 34,000 and 30,000 mol wt IGFBPs, which may transport IGF-I to tissues. The faster serum elimination half-life may result from diminished IGFBP-3, leading to greater IGF-I availability to tissues in pregnancy.

Animals↗

PubMed1990

Insulin-like growth factor binding protein secretion by breast carcinoma cell lines: correlation with estrogen receptor status.

Breast tumor cell lines have been shown to secrete several distinct polypeptide growth factors, although conflicting results exist for the insulin-like growth factors (IGFs). In contrast a limited number of breast tumor cell lines have definitely been shown to secrete the high affinity IGF binding proteins (IGFBPs) that modify IGF actions. To characterize the types of IGFBPs that are secreted by breast tumor cell lines, conditioned medium was collected from seven separate tumor cell lines, three of which were estrogen receptor (ER) negative, and four of which were ER positive. All three of the ER negative cell lines, MDA-231, MDA-330, and HS578T, secreted binding proteins of 49,000 and 43,000 Mr (IGFBP-3) as well as 29,000 (IGFBP-1) and 24,000 Mr. In contrast, all four ER positive cell lines secreted 34,000 (IGFBP-2) or 24,000 Mr forms, and none secreted the 49,000 and 43,000 or 29,000 Mr forms. BT-20, a cell line that is positive for ER messenger RNA (mRNA) but negative for ER protein, secreted predominantly a 34,000 Mr protein. The amount of total IGFBP activity released in 24 h ranged between 0.4 and 5.6 nM equivalents of IGFBP-1, and there was no significant difference between the ER positive and negative cell lines. The MCF-7 cells that produced predominantly 34,000 and 24,000 Mr forms showed a 1.8-fold increase in IGFBP secretion after estrogen stimulation. Immunoblotting and a specific RIA for IGFBP-1 showed that only the ER negative lines MDA-330, MDA-231, and HS578T secreted this form. Northern blotting analysis for the mRNA encoding this protein showed that both MDA-330 and MDA-231 contained a single 1.6 kilobase mRNA species that hybridized with an IGFBP-1 complementary DNA (cDNA) probe. Immunoblotting analysis of the other cell lines showed that only the 34,000 Mr form secreted by the ER positive cell lines reacted with IGFBP-2 antisera. Exposure of the conditioned media from the three ER negative cell lines to N-glycanase revealed that the 49,000 and 43,000 Mr forms of IGFBP were glycosylated and therefore probably represent IGFBP-3. We conclude that ER negative cell lines secrete three forms of IGFBPs, IGFBP-1, IGFBP-3, and a 24,000 Mr form. In contrast, the ER positive cell lines secrete predominantly IGFBP-2 and the 24,000 Mr form but do not secrete IGFBP-3 or 1.(ABSTRACT TRUNCATED AT 400 WORDS)

Biomarkers, Tumor↗

PubMed1990

Insulin-dependent regulation of insulin-like growth factor-binding protein-1.

In previous studies it has been determined that food ingestion or glucose infusion suppresses serum insulin-like growth factor-binding protein-1 (IGFBP-1) concentrations in normal subjects. It has not been determined, however, whether glucose-dependent suppression is due to enhancement of insulin-stimulated glucose transport, to some other insulin-mediated effect, or to a direct effect of glucose. These studies were undertaken to determine if infusion of other nutrients or energy sources that activate glycolysis, but not insulin secretion, could lead to suppression of the plasma concentrations of this protein. After infusion of 50 g glucose over 4 h, the mean plasma IGFBP-1 concentration fell from 44.3 +/- 13.8 to 17.4 +/- 8.1 micrograms/L (P less than 0.001). In contrast, when 50 g fructose were infused over 4 h, IGFBP-1 decreased from 44.6 +/- 7.4 to only 32.7 +/- 6.7 micrograms/L (P less than 0.01). Comparison of these changes showed that the mean decrease after glucose infusion was significantly greater than that after fructose (P less than 0.01). In contrast to these decreases, infusion of an isocaloric amount of triglycerides resulted in no significant change in IGFBP-1 concentrations (from 49.8 +/- 14.1 to 40.2 + 10.4 micrograms/L; P = NS). Concomitant measurements of immunoreactive C-peptide during the glucose and fructose infusions showed that plasma C-peptide levels rose from 1.6 +/- 0.2 to 3.9 +/- 0.7 nM (P less than 0.001) during the glucose infusion, whereas the maximum increase was from 1.7 +/- 0.4 to only 2.3 +/- 0.4 nM (P less than 0.05) during the fructose infusion. The difference between these mean changes was also significant. The change in C-peptide correlated inversely with the changes in IGFBP-1 during the glucose infusion (r = -0.68); P less than 0.001), but not during fructose infusion (r = -0.31). The results of this study suggest that insulin stimulation of cellular metabolic pathways is an important variable regulating the suppression of plasma IGFBP-1 concentrations. Activation of noninsulin-dependent glycolytic pathways appears to result in an equivalent degree of suppression, whereas provision of other energy substrates has no effect. We conclude that both insulin and glucose are important regulators of plasma concentrations of IGFBP-1 in vivo, and in this way they may significantly influence IGF action.

Adult↗

PubMed1989

Insulin-like growth factors (IGF) in muscle development. Expression of IGF-I, the IGF-I receptor, and an IGF binding protein during myoblast differentiation.

The insulin-like growth factors (IGFs) I and II exert pleiotropic effects on diverse cell types through interaction with specific high affinity cell surface receptors and with locally produced binding proteins. In skeletal muscle and in myoblast cell lines, the functions of IGF-I and -II are complex. Both growth factors appear capable of stimulating cellular proliferation and differentiation, as well as exerting insulin-like effects on intermediary metabolism. We have demonstrated recently that the expression of IGF-II and its receptor is induced during the terminal differentiation of the myoblast cell line, C2, and have suggested that IGF-II may be an autocrine growth factor in these cells (Tollefsen, S.E., Sadow, J.L., and Rotwein, P. (1989) Proc. Natl. Acad. Sci. U.S.A. 86, 1543-1547). We now have examined this cell line for expression of other components involved in IGF signaling. The synthesis of IGF-I is low during myoblast proliferation; IGF-I mRNA can be detected only through use of a sensitive solution hybridization assay. Typical IGF-I receptors can be measured in myoblasts, whereas IGF binding proteins cannot be detected in proliferating cells or in conditioned culture medium. During myogenic differentiation, IGF-I mRNA levels increase transiently by 6-10-fold within 48-72 h. The expression of IGF-I mRNA is accompanied by a 2.5-fold accumulation of IGF-I in the culture medium. IGF-I receptors also increase transiently, doubling by 48 h after the onset of differentiation. By contrast, secretion of a Mr 29,000 IGF binding protein is induced 30-fold to 100 ng/ml within 16 h and continues to increase throughout differentiation. These studies demonstrate that several components critical to IGF action are produced in a fusing skeletal muscle cell line in a differentiation-dependent manner and suggest that both IGF-I and IGF-II may be autocrine factors for muscle.

Amino Acid Sequence↗