Somatomedin-binding Protein Research Articles

Renal Hemodynamics in Recent-Onset Type II Diabetes

Glomerular hyperfiltration is thought to play an important role in the genesis of diabetic nephropathy. While hyperfiltration is well documented in early type I diabetes, the evidence for hyperfiltration in type II diabetes is conflicting. We investigated 16 nonproteinuric patients with recently diagnosed type II diabetes. Glomerular filtration rate (GFR) and effective renal plasma flow (ERPF) were measured as inulin clearance (CIN) and p-aminohippuric acid clearance (CPAH) using a constant infusion technique. Lean body mass was measured by densitometry (weighing under water). Renal hemodynamics were also measured in 31 healthy volunteers and six obese nondiabetic individuals. Median GFR in diabetics (133 mL/min/1.73 m2; range, 95 to 165) was significantly (P < 0.01) higher than in obese nondiabetic controls (median, 118; range, 95 to 139). Elevated GFR (ie, >95th percentile of non obese healthy controls) was found in 44% of patients. When GFR was factored for lean body mass, it was elevated in 50%. GFR did not correlate with fasting glucose, hemoglobin A1c (HbA1c), insulin-like growth factors, IGF-1 and IGF-2, or somatomedin-binding protein (SMBP). The findings document that hyperfiltration is common in recent-onset type II diabetics.

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 (E 2)-induced growth suggesting that secreted IGFs are not the major mediators of E 2 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 ≅ 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 the IGFs are important regulators of breast cancer cell proliferation and that antagonism of this pathway may offer a new treatment strategy.

A Specific Radioimmunoassay for the Growth Hormone (GH)-Dependent Somatomedin-Binding Protein: Its Use for Diagnosis of GH Deficiency

The acid-stable subunit of the GH-dependent large mol wt somatomedin-binding protein (SmBP) was isolated from human plasma Cohn fraction IV by a three-step procedure, and a specific RIA was developed which allowed measurement in unextracted serum. Although in normal human serum most of immunoreactive material was present as the large mol wt complex (150K), considerable amounts of smaller components were found by high performance liquid exclusion chromatography in the 60K, 42K, and 32K range. Normal serum levels were low at birth, rose sharply during the first weeks of life, and showed a moderate peak at puberty. To assess the diagnostic efficacy of SmBP for GH deficiency (GHD), patients previously diagnosed as GH-deficient by conventional criteria (n = 132) were compared to short statured children without GHD (n = 130). Taking the fifth centile as a limit of normality the majority of patients with GHD had subnormal levels, yielding high sensitivity of the test (0.97). In contrast, most of the non-GH-deficient children had SmBP levels within the normal range, resulting in high specificity (0.95) and, consequently, high accuracy (0.96). These results suggest that the large mol wt SmBP is an excellent screening parameter and is highly informative for GHD.

Growth Hormone Induced Rise in Glomerular Filtration Rate Is Not Obliterated by Angiotensin-Converting Enzyme Inhibitors

In 8 healthy normotensive probands with normal glomerular filtration rate, the effect of recombinant human growth hormone (rhGH) on inulin clearance (Cin) was examined in an open study with intraindividual crossover with or without enalapril pretreatment (20 mg/day). rhGH was administered by subcutaneous injection (4.5 U twice daly) for 3 days. On the following day Cin was measured with an enzymatic steady state infusion technique. Systemic hemodynamics and potential metabolic effects of rhGH, i.e., inulin-like growth factors I and II, somatomedin-binding protein, glucagon, C peptide, amino acid pattern, etc., were monitored. On controlled dietary intake of protein, the median Cin rose 72 h after start of rhGh administration from 114 (range 91-158) to 135 (108-167) ml/min/1.73 m2 without enalapril pretreatment (p less than 0.01) and from 111 (88-153) to 131 (100-173 ml/min/1.73 m2 with enalapril pretreatment (p less than 0.02). The results confirm that (1) rhGH increases Cin to a similar extent as extractive GH and (2) further demonstrate that this action is not obliterated by blocking the circulating converting enzyme.

Studies on Insulin-like Growth Factors (IGF-I, -II) and There Binding Proteins in Normal Human Pregnancy

In order to elucidate the roles of Insulin-like Growth Factors (IGF-I, -II) in human pregnancy, the levels of IGF-I and IGF-II, the distribution of binding proteins for IGF-I or IGF-II and profiles of unsaturated somatomedin binding proteins (USBP) were estimated in maternal and cord plasma. IGF-I levels in maternal plasma gradually elevated in late gestation, reaching 304.4 +/- 130.1ng/ml at term, and were significantly higher than those in nonpregnant women (188 +/- 58). IGF-II levels, which were 414.9 +/- 75.4ng/ml in women in the third trimester of gestation, did not produce any remarkable changes in the levels in nonpregnant women (395.9 +/- 72.6). On the other hand, IGF-I in cord plasma also increased according to gestational age, reaching 37.3 +/- 14.6ng/ml at term, but it was significantly lower than that in maternal weight (r = 0.50, p less than 0.005) and relative birth weight (r = 0.41, p less than 0.005). IGF-II in cord plasma showed no significant changes during gestation, however, IGF-II levels in AFD (appropriate for date) newborns delivered at term (203.8 +/- 59.4) were significantly lower than those in maternal plasma. And they had no positive correlations with birth weight and relative birth weight. On Sephadex G150 gel-chromatography of cord plasma from AFD newborns at term, more than 70% of immunoreactive IGF-I in plasma was eluted at 150K region, and 150K USBP could be detected as observed in adult plasma. On the other hand, most of the immunoreactive IGF-II was eluted at 40K region, and 150K USBP was not detected in AFD newborns at term. In adult plasma, most of the immunoreactive IGF-II was eluted at 150K region, but 150K USBP could not be detected. From these results, it is supposed that IGF-I plays an essential role in fetal growth rather than IGF-II.

Somatomedin-binding proteins: What role do they play in the growth process?

The insulin-like growth factors (IGF-I and IGF-II) circulate bound to specific proteins. Two classes of binding proteins have been relatively well characterized differing in size, immunological activity, and physiological function. This review summarizes the more recent data on IGF-binding proteins, focusing on their possible clinical implications.

Somatomedin binding protein and somatomedin C are reliable indicators of acromegalic activity

Somatomedin binding protein and somatomedin C are reliable indicators of acromegalic activity

The value of IGF-I, IGF-II, and somatomedin binding protein for the diagnosis of growth hormone deficiency

The value of IGF-I, IGF-II, and somatomedin binding protein for the diagnosis of growth hormone deficiency

Circulating Forms of Insulin-like Growth Factor-I (IGF-I) /Somatomedin C (SMC) in Fetal Life

IGF-I/SMC shows mitogenic activities in a wide variety of cell types and stimulates cell growth in vitro and in vivo. Unlike most other peptide hormones, IGF-I/SMC circulates in the plasma as macromolecular complexes with specific plasma binding proteins, approximate molecular weight of 150K and 40K daltons. In order to elucidate the roles of IGF-I/SMC for fetal and postnatal development, the levels of plasma IGF-I/SMC, distribution of circulating IGF-I/SMC on its binding proteins, and profiles of unsaturated somatomedin binding proteins (USBP) were estimated in newborns and a growth-retarded infant (leprechaunism). The concentrations of IGF-I/SMC in cord plasma increased gradually after the 28th week of gestation and reached 37.3 +/- 14.6 ng/ml (Mean +/- S.D.) in full term infants. Although the levels of IGF-I/SMC in cord plasma were significantly lower than those in non-pregnant women (188 +/- 58), they showed a positive correlation with birth weight and relative birth weight (R.B.W.: percentage comparison to the average birth weight of normal infants in the same gestational week). In adult plasma, immunoreactive IGF-I/SMC was eluted predominantly in 150K dalton region (73.5%) and to a lesser extent (26.5%) in 40K dalton, and two apparent peaks of USBP could also be determined in both 150K and 40K dalton regions. On the other hand, in fetal plasma 84.4% of immunoreactive IGF-I/SMC was eluted in 40K region on the 20th week, but on the 26th week, 71.6% of IGF-I/SMC was eluted in 150K region and after the 35th week, more than 70% of IGF-I/SMC was determined in 150K region as observed in adult plasma. However, 150K.USBP could not be detected in cord plasma obtained from appropriate-for-date(AFD) infants until after the 35th week of gestation. Additionally, a positive correlation was demonstrated between 150K.USBP/40K.USBP ratio and birth weight. Some cases of small-for-date(SFD) infants whose IGF-I/SMC circulated mainly as 150K complex could catch up on their growth early in postnatal life. However, in the SFD newborns, if either 150K.USBP or 150Kcomplex of IGF-I/SMC could not be detected, their growth spurt was remarkably delayed until a year after birth. Furthermore, the similar disorders of IGF-I/SMC distribution and its USBP profile were also observed in the case of leprechaunism, who showed severe intrauterine and postnatal growth retardation.(ABSTRACT TRUNCATED AT 400 WORDS)

A Case of Leprechaunism with the Disorders of Insulin-like Growth Factor-I (IGF-I) /Somatomedin C (SMC) Binding Protein and Its Receptor

Leprechaunism is a rare genetic disorder characterized by physical abnormalities, intrauterine and postnatal growth retardation, poorly developed subcutaneous fat and muscle at birth, and early death. This patient, who was a 1.5 year-old female with typical clinical features of leprechaunism, had relatively high levels of plasma GH and IGF-I/SMC but no glucose intolerance or insulin resistance. Studies were undertaken to elucidate (1) the differences among some kinds of methods for IGF-I/SMC measurement, (2) the distribution patterns of IGF-I/SMC between two kinds of its binding protein (SMBP) in plasma, and (3) the dynamics of IGF-I/SMC receptor in her erythrocytes and liver microsomal membranes. The results were as follows: (1) The level of IGF-I/SMC measured by Nichols radioimmunoassay kit was 1.33U/ml, which was higher than that of infants the same age. Conversely, it was lower than that of the control which was measured by radioimmunoassay using recombinant IGF-I/SMC after acid-ethanol or Seppak C18 extraction. (2) By Sephadex G150 gel-chromatography, immunoreactive IGF-I/SMC was eluted predominantly in 150K region, and two apparent peaks of unsaturated somatomedin binding protein (USBP) were determined in a neonatal infant (appropriate to date), a normal adult and an infant of the same age as this patient. On the other hand, immunoreactive IGF-I/SMC was located only in the fractions corresponding to 40K region, and only one peak of USBP could be estimated in the region of 40K dalton. (3) The IGF-I/SMC receptor in the patient's erythrocytes possessed significantly lower binding affinity but higher binding capacity in comparison with that of the normal neonate and adult. In addition, the receptor in liver microsomal membranes obtained from this patient at autopsy also indicated lower affinity but higher capacity than that of fetuses at more than 19 weeks of gestation. This was coincident to that of fetuses less than 19 weeks of gestation. These results suggested that this patient resembled the intrauterine fetus before midgestation not only in the co-relationship among GH, IGF-I/SMC and its binding proteins, but also in the characteristics of its receptor. The severe growth retardation existing in this patient may be, at least partly, due to the abnormality and/or immaturity of IGF-I/SMC function. It is speculated that leprechaunism could be classified in relation to fetal growth mechanism by aspects of biological functions of IGF-I/SMC during development.

Discordance of serum and tissue somatomedin levels in growth hormone-stimulated growth in the rat.

Although the somatomedins are believed to mediate GH-induced somatic growth, circulating levels of insulin-like growth factor I (IGF-I) do not always correlate with growth rate. To evaluate a paracrine or autocrine effect that might explain this discordance, the concentrations of plasma and tissue IGF-I were compared with indices of growth in hypophysectomized rats treated with rat GH (rGH). The rats received a total of 250 micrograms rGH by either continuous pump infusion (P) or twice daily injections (I). After 4 days of treatment, the amounts of IGF-I present in acetic acid extracts of liver and kidney and in native serum were determined by RIA and related to proximal tibial epiphyseal plate width. Tibial epiphyseal plate width increased from 198 +/- 6 microns (mean +/- SE) in untreated controls to 339 +/- 13 microns in group P and 347 +/- 9 micron in group I; weight gain was 11.6 +/- 1.0 g in group P and 9.6 +/- 0.8 g in group I, while control animals lost 1.0 g. Serum concentrations of IGF-I were no different between control animals and those in the injection group (0.91 +/- 0.06 vs. 0.90 +/- 0.06 U/ml), whereas levels in the infusion group increased slightly (1.1 +/- 0.05 U/ml). In contrast, rGH administration caused tissue IGF-I to double in liver (control, 0.24 +/- 0.04 U/g; P, 0.51 +/- 0.03 U/g; I, 0.46 +/- 0.05 U/g) and nearly triple in kidney (control, 0.52 +/- 0.05 U/g; P, 1.51 +/- 0.09 U/g; I, 1.36 +/- 0.08 U/g). There was no detectable change in somatomedin-binding protein by gel exclusion chromatography. Since the rGH administered was sufficient to stimulate growth and increase tissue somatomedin levels without corresponding increases in circulating IGF-I, an autocrine or paracrine action of IGF-I appears to mediate GH's initial somatogenic actions in the young rat.

Influence of parasitism on plasma concentrations of growth hormone, somatomedin-C and somatomedin-binding proteins in calves

A parasitic disease model (sarcocystosis) was used to study the effects of infection and associated plane of nutrition on GH and somatomedin-C (SM-C) patterns in plasma, and SM-C binding protein patterns in plasma from 4-month-old male Holstein calves. Calves, matched by age and rate of growth before the experiment, were divided into three treatment groups (n = 7). In the first (control), animals were uninfected and food was available ad libitum; in the second, animals were infected with Sarcocystis cruzi and food was available ad libitum. The third group consisted of uninfected animals pair-fed to the level of feed intake of the infected animals. Blood samples were obtained at various times after infection for analysis of the secretory patterns of GH (day 27 after infection, samples every 10 min for 6 h), SM-C (days 27, 35 and 58 after infection) or binding protein (day 42 after infection). Samples were analysed for GH and SM-C by radioimmunoassay. Relative molecular weights of binding proteins were assessed by elution patterns from gel permeation columns. Clinical signs of infection were manifest abruptly on day 26 after infection. Voluntary feed intakes of infected calves as a per cent of control calves were 18, 46 and 78 on days 27, 35 and 58 after infection respectively. Plasma GH concentrations were lower in infected and pair-fed than in control calves (P less than 0.05). Plasma SM-C concentrations were reduced in calves with diminished feed intakes and lower still in infected calves (P less than 0.05). Plasma SM-C was positively correlated with nitrogen retention across treatment groups (r = 0.81). Two classes of binding proteins differing in molecular weight were identified. The relative amounts of each binding protein in plasma were reduced during low feed intake with some differences in the endogenous saturation affected by infection. These data suggest that altered growth and metabolism in parasitized calves may arise in part from both nutritional and infection-mediated effects on the regulation of GH, SM-C and SM-C binding proteins.

Clinical Significance of Somtomedins and Somatomedin Binding Proteins

Clinical Significance of Somtomedins and Somatomedin Binding Proteins

Cloning of cDNA encoding human placental protein 12 (PP12): binding protein for IGF I and somatomedin.

By N-terminal sequence comparison it was shown that the placental protein 12 (PP12) is homologous to the IGF I (insulin-like growth factor) and somatomedin binding protein. Starting with an antibody raised against PP12 the authors have screened cDNA libraries from human placenta. Two cDNA clones were identified with an open reading frame encoding a hydrophobic leader peptide consisting of 25 aa and the mature protein of 234 aa (MW 25 270). The MW and the amino acids composition are in good agreement to the data of Bohn and Kraus. The protein sequence shows no N-linked glycosylation site. The cDNA encodes 18 Cys-residues, which may explain the wide discrepancy in the reported MW of the PP12 protein. The stop codon at position 928 is followed by 566 nt of untranslated region including the poly(A) addition site AATAAA 18 nt upstream of poly(A).

Clinical Significance of Somtomedins and Somatomedin Binding Proteins

Clinical Significance of Somtomedins and Somatomedin Binding Proteins

Time-resolved immunofluorometric assay of 34-kDa somatomedin-binding protein.

In this time-resolved immunofluorometric assay for the 34-kDa somatomedin-binding protein (SmBP), affinity-purified polyclonal antibodies are used, along with solid-phase separation of bound and free analyte. The first antibody is bound to polystyrene microtiter wells; the second is labeled with europium(III) chelate. The detection limit of the method is 0.25 microgram/L, much lower than that (about 8 micrograms/L) for radioimmunoassay. By immunofluorometric assay, SmBP is detectable, and could be accurately quantified, in the serum of all 88 individuals we tested, whereas by radioimmunoassay a third of the samples had concentrations below the detection limit. When SmBP was detectable by both methods, the concentrations measured by the two techniques correlated well (r = 0.98).

Purification of placental protein PP12 from human amniotic fluid and its comparison with PP12 from placenta by immunological, physicochemical and somatomedin-binding properties

Human amniotic fluid was found to contain a protein which is immunochemically indistinguishable from placental protein PP12. This protein was purified by gel filtration, hydrophobic interaction high performance liquid chromatography and anion-exchange chromatography. The relative molecular mass as determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis was 34000, and the isoelectric point was 4.9. Tryptic peptides of amniotic fluid PP12 as determined by reversed phase high performance liquid chromatography were similar to those of placental PP12. Both had the N-terminal amino acid sequence Ala-Pro-Trp-Gln-, which is the same as previously reported for a somatomedin-binding protein. Both placental PP12 and amniotic fluid PP12 were found to bind somatomedin C (IGF-I) with high affinity, K a = 1 × 10 9 l/mol). Amniotic fluid is an ideal source of this somatomedin-binding protein, and the purification method described allows rapid isolation of PP12 under mild conditions which are essential for studies on its biological function.

Serum somatomedin binding proteins: physiologic significance and interference in radioligand assay.

Mammalian sera contain binding proteins that specifically complex with somatomedins (insulin-like growth factor I and II) so that there are no detectable free somatomedins. There is immunologic evidence of two distinct types of serum binding proteins. The major binding protein complex (150,000 Mr) is growth hormone dependent. Binding proteins protect somatomedins from proteolytic degradation, retard plasma clearance, and decrease the availability to tissue receptors. The presence of serum binding proteins interferes with radioimmunoassays and radioreceptor assays for somatomedins. Proposed strategies to neutralize the interference from binding proteins without their elimination do not achieve this goal. Extraction of somatomedins by acid ethanol, hydrophobic absorption on C18 silicates (SepPak), and acid gel filtration are effective with human serum, but only acid gel filtration is satisfactory with rat serum. Failure to eliminate binding proteins from assays can lead to serious artifacts in conditions where abnormalities of binding proteins exist.

Synthesis and Circulating Forms of The Somatomedins

Our data support the hypothesis that the somatomedins have multiple origins and may have paracrine and autocrine as well as humoral modes of action. The balance between the local actions and hormoral actions of somatomedin in any physiologic circumstance remains to be determined by future experiments. We have also found evidence that the initial products of IGF secretion synthesis include a carboxyterminal E region extension as predicted by the messenger RNA structure. In normal circumstances, the E region appears to be rapidly and efficiently cleaved. However, in certain metabolic conditions IGF-I prohormone is found in the circulation. Our initial characterization of the circulating IGF-I prohormone suggests that the prohormone form of IGF is unable to bind to either the type IGF receptor or to the binding proteins. In addition, we have demonstrated that normal plasma contains an active and specific peptidase for the E region, which is associated with molecules of the 150, 000 molecular weight range in plasma. A small binding protein for somatomedin has been isolated from a number of human sources and partial sequence data has been reported. There is at least some immunological and structural homology between the binding protein associated with the 150, 000 micro-weight complex and the small somatomedin binding protein. Biological regulation of somatomedin synthesis and action can occur at multiple sites, including messenger RNA synthesis, prohormone synthesis and processing, and incorporation into mature somatomedin complexes in plasma. The biological control of these processes, and their relationship to the biological actions of the somatomedins, will be a source of fruitful future study.

Progesterone-associated proteins PP12 and PP14 in the human endometrium

Two proteins, designated as PP12 and PP14 were originally isolated from soluble extracts of the human placenta and its adjacent membranes. We have shown that they are synthesized by decidualized/secretory endometrium and not by placenta. Both proteins occur at high concentrations in human amniotic fluid, which is therefore an excellent source for purification. PP12 is a 34-kDa glycoprotein, which has an N-terminal amino acid sequence of Ala-Pro-Trp-Gln-Cys-Ala-Pro-Cys-Ser-Ala. This is identical with that of somatomedin-binding protein purified from the amniotic fluid. PP12 too binds somatomedin-C, or IGF-I (insulin-like growth factor-I). Human secretory endometrium synthesizes and secretes PP12, and progesterone stimulates its secretion. PP14 is a 28-kDa glycoprotein. Its N-terminal sequence shows homology to that of beta-lactoglobulins from various species. We have found PP14 in the human endometrium, serum and milk. Immunologically, PP14 is related to progestagenassociated endometrial protein (PEP), alpha-2 pregnancy-associated endometrial protein (alpha-2, PEG), endometrial protein 15 (EP15), alpha-uterine protein (AUP) and chorionic alpha-2 microglobulin (CAG-2). In ovulatory menstrual cycles, the concentration of PP14 increases in endometrial tissue as the secretory changes advance. In serum, the PP14 concentration begins to rise later than the progesterone levels, and hih serum PP14 levels are maintained for the first days of the next cycle. By contrast, no elevation of serum PP14 level is seen in anovulatory cycles. Our results show that progesterone-associated proteins are synthesized by the human endometrium and appear in the peripheral circulation, where they can be quantitatively measured using immunochemical techniques.