Fertility and Pregnancy in Primary Adrenal Insufficiency in Germany.

Autoimmune adrenalitis (AA) and congenital adrenal hyperplasia (CAH) are the most common reasons for acquired and monogenetic primary adrenal insufficiency. Both concern women in their fertile years. The aim of the work was to examine fertility rates, pregnancy outcome, and children's characteristics in AA and CAH patients in 2 German endocrine centers. One hundred and fifty-eight women were contacted. Thirty-nine patients with CAH due to 21-hydroxlase deficiency and 54 AA patients agreed and were included. Information about course and outcome of pregnancies was obtained by questionnaire and telephone interview. Fertility rates were calculated and compared to expected values from the German general population. Twelve CAH patients (30.8%) had 25 pregnancies, which resulted in 16 children. In AA patients, 93 pregnancies in 42 women (75%) were reported resulting in 73 childbirths. Fertility rates were normal in nonclassic CAH and in AA-only patients, but significantly reduced in classic CAH and autoimmune polyendocrine syndrome type 2 (APS-2). Rates of miscarriages were high in all CAH (36%) and APS-2 (22%) patients. The majority of children in both groups were born at term, but rates of cesarean section were elevated in classic CAH and in AA patients<5 years before or after diagnosis. Children born to CAH patients weighed significantly less than expected and 33.3% of them were born small for gestational age. Fertility seems not to be reduced in general, but specific in classic CAH and APS 2 patients. Nevertheless all CAH and AA patients seem to be at risk of miscarriages and cesarean section.

High-resolution record of Northern Hemisphere climate extending into the last interglacial period.

Two deep ice cores from central Greenland, drilled in the 1990s, have played a key role in climate reconstructions of the Northern Hemisphere, but the oldest sections of the cores were disturbed in chronology owing to ice folding near the bedrock. Here we present an undisturbed climate record from a North Greenland ice core, which extends back to 123,000 years before the present, within the last interglacial period. The oxygen isotopes in the ice imply that climate was stable during the last interglacial period, with temperatures 5 degrees C warmer than today. We find unexpectedly large temperature differences between our new record from northern Greenland and the undisturbed sections of the cores from central Greenland, suggesting that the extent of ice in the Northern Hemisphere modulated the latitudinal temperature gradients in Greenland. This record shows a slow decline in temperatures that marked the initiation of the last glacial period. Our record reveals a hitherto unrecognized warm period initiated by an abrupt climate warming about 115,000 years ago, before glacial conditions were fully developed. This event does not appear to have an immediate Antarctic counterpart, suggesting that the climate see-saw between the hemispheres (which dominated the last glacial period) was not operating at this time.

Expression of insulinlike growth factor (IGF) and IGF-binding protein genes in human lung tumor cell lines.

BACKGROUND: The presence of multiple, low-molecular-weight, insulinlike growth factor (IGF)-binding proteins in lung tumor cell-conditioned medium and lung cancer patient serum has been recently reported. PURPOSE: To begin to elucidate the genetic basis for these observations, the present study examines the expression by lung tumor cell lines of three IGF-binding protein genes, namely, IGFBP-1, IGFBP-2, and IGFBP-3. Since IGF-binding proteins are thought to modulate the biologic action of the IGFs, the relationship between the expression of IGF-binding protein genes and the genes encoding IGF-I and IGF-II also has been investigated. METHODS: Gene expression was studied in four small-cell lung cancer (SCLC) and three non-small-cell lung cancer (NSCLC) cell lines using Northern blot analysis and reverse transcriptase polymerase chain reaction (RT-PCR) for IGFBP-1. RESULTS: IGFBP-1 gene expression was detected by Northern blot analysis in one NSCLC cell line only. However, RT-PCR revealed that the IGFBP-1 gene was expressed in all four SCLC cell lines and in two of the three NSCLC lines. Northern blot analysis of IGFBP-2 gene expression demonstrated that all lung tumor cell lines expressed this gene. A low level of IGFBP-3 gene expression was detected in one SCLC cell line and in all three NSCLC cell lines. All lung tumor cell lines expressed the IGF-II gene as determined by Northern blot analysis. In marked contrast, none of the lines showed evidence of IGF-I gene expression using this method. However, RT-PCR revealed a low level of IGF-I gene expression in one SCLC and one NSCLC cell line only. CONCLUSIONS: These observations indicate 1) that IGF-binding proteins secreted by lung tumors are encoded by at least three different genes; 2) that there may be a close association between IGF-II and IGFBP-2 gene expression, such that, where there is production of IGF-II, IGFBP-2 is the principal BP; and 3) that the IGF-II gene is more widely expressed than the IGF-I gene in human lung tumor cell lines.

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

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

Genomic structure and regulation of the promoter of the rat insulin-like growth factor binding protein-2 gene.

We describe the complete genomic organization of the rat insulin-like growth factor binding protein-2 (rIGFBP-2) gene. This single-copy gene spans over 36 kilobases (kb) and is split into four exons of 475, 224, 141, and 472 nucleotides (nt), and three introns of 32 kb, 686, and 1793 nt, respectively. A single transcription start site (-90) was mapped by S1 protection assay and primer extension. The putative promoter of the rIGFBP-2 gene does not possess TATA or CAAT elements; however, it contains three GC-rich regions located 37, 57, and 81 nt 5' of the cap site. Deletion analysis of the 0.6-kb region of the upstream sequences and transfection of these constructs into BRL-3A and Chinese hamster ovary cells were used to localize possible cis-acting elements. The three GC boxes enhanced chloramphenicol acetyltransferase and luciferase transcription almost to the same level as the XbaI-NsphI (-579 to +1) fragment and displayed synergism and orientation dependence. In addition a similar positive effect on luciferase transcription has been obtained by cotransfecting these fragments with varying amounts of Sp1 expression vector into Drosophila cells that lack endogenous Sp1. In vitro gel mobility shift assays demonstrated that box 1 (GGGCGG), box 2 (GGGAGG), and box 3 (GGGAAGG) bind to SpI with variable affinities and display cooperativity. A protein that gave a similar DNA binding pattern was present in nuclear extracts of BRL-3A cells. To analysis using consensus or aberrant Sp1 elements and a polyclonal Sp1 antiserum to inhibit DNA binding were performed. These in vivo and in vitro data demonstrated that Sp1 plays an important role in the regulation of the expression of rIGFBP-2.

Structure of the human insulin-like growth factor binding protein-2 gene.

Insulin-like growth factors (IGFs) are polypeptide hormones with structural homology to proinsulin. IGFs circulate in blood bound to specific IGF binding proteins (IGFBPs). cDNA sequences of six members of a family of human and rat IGFBPs have been published. Here we present a partial characterization of the human IGFBP-2 gene. This single copy gene is located on chromosome 2 and spans a total of more than 32 kilobases (kb) of genomic sequence. It is organized in four exons with sizes of more than 568, 220, 141, and 496 nucleotides. The intron between exon one and exon two contributes 27 kb to the size of the IGFBP-2 gene. The second and the third introns comprise 1.1 kb and 1.95 kb, respectively. When the structure of the IGFBP-2 gene is compared to that of the IGFBP-1 and IGFBP-3 genes, the exon boundaries are found to be conserved in these three genes. A single transcriptional start site was localized to 113 +/- 2 nucleotides 5' of the ATG start codon of IGFBP-2 translation. Furthermore, the region between nucleotides -635 and -2 upstream of the ATG was demonstrated to exhibit promoter activity in human Jurkat K16 cells. This region is devoid of TATA or CAAT consensus sequence motifs and has a high content of dC and dG nucleotides. In this respect the putative IGFBP-2 promoter region resembles the promoters which are often associated with housekeeping genes.

A low molecular weight insulin-like growth factor binding protein from rat: cDNA cloning and tissue distribution of its messenger RNA.

Rat serum contains two major forms of insulin-like growth factor (IGF) binding proteins (BPs) that have apparent mol wts of about 35,000 and 150,000. We have isolated a cDNA clone encoding an IGF-BP whose N-terminal sequence is completely homologous to the NH2-terminal of the Buffalo rat liver cells-3A BP. The 270 amino acid mature protein has a predicted mol wt of 29,500. It contains a cysteine rich domain at each end of the molecule and an Arg-Gly-Asp (RGD) tripeptide motif near its C-terminus which suggests that this BP might associate with integrin cell surface receptors. The mature protein shares only partial homology with two published human IGF-BPs. Northern blot analysis shows that its mRNA is abundant in several fetal tissues, in adult brain, testes, ovaries, and kidney. Expression in the liver is high in fetal life but decreases to a barely detectable level in adulthood. However, upon hypophysectomy, the mRNA level increases at least 20-fold which suggests a hormonal regulation for the hepatic production of this small IGF-BP.

Insulin regulates the expression of the insulin-like growth factor binding protein 2 mRNA in rat hepatocytes.

The goal of this study was to find out whether GH or insulin regulate the mRNA expression of the fetal binding protein of insulin-like growth factor (IGFBP-2). Primary hepatocytes from adult rats were used as a test system. IGFBP-2 mRNA was abundant in cells cultured in the absence of hormones and markedly reduced in cultures containing insulin. Addition of GH had no effect on IGFBP-2 mRNA levels although the cells are responsive to GH as demonstrated by a GH mediated elevation of IGF l mRNA levels. Half-maximal down-regulation of IGFBP-2 mRNA levels occurred at an insulin concentration of 1 to 2 x 10(-10) M. The finding that insulin is a potent negative regulator of hepatic IGFBP-2 mRNA levels suggests a physiologically important regulatory link between the two hormones insulin and IGF l.

Synthesis and secretion of insulin-like growth factor and its binding protein by the perfused rat liver: dependence on growth hormone status.

Isolated livers of normal and hypophysectomized (hypox) rats with or without GH replacement therapy were perfused in an erythrocyte-free recirculating perfusion system for 4 h in the presence of [35S]cysteine. Albumin secretion and synthesis increased in a parallel and linear fashion over 4 h. The albumin secretion rates were 0.53 and 0.21 mg/g liver h-1 in normal and hypox animals, respectively. Insulin-like growth factor (IGF) secretion, measured as insulin equivalents in the fat cell assay as well as in a competitive protein binding assay, and IGF synthesis, as determined from [35S]cysteine incorporation into immunoprecipitable IGF, likewise increased linearly and in parallel throughout the perfusion time. The IGF secretion rate was 50 microU/g liver h-1. The secreted IGF had a molecular weight of approximately 7700 daltons. Secretion and synthesis of IGF were reduced to 11% in hypox rats and were largely restored by human GH replacement therapy (to 86% of normal). A single specific binding protein with an approximate molecular weight of 35,000 was detected in the perfusate. The binding protein was measured by covalent cross-linkage to [125I]IGF I by dimethylsuberimidate. The secretion of this binding protein was 62% of normal in hypox animals and 79% in GH-treated hypox rats. The data suggest that IGF is continuously synthesized and released by the liver. Assuming a half-life for IGF of 3 h in the normal rat, a plasma volume of 8 ml, and a liver weight of 8.5 g, the rate of IGF production by the perfused normal rat liver (50 microU/g liver h-1) would be sufficient to maintain serum IGF at the concentration determined in normal rat serum (approximately 130 microU/ml). This suggests that the liver is the major site of IGF production in the rat.

Expression of the BRL-3A insulin-like growth factor binding protein (rBP-30) in the rat central nervous system.

The expression of the BRL 3A insulin-like growth factor binding protein (rBP-30) was characterized in rat brain, hypothalamus, and pituitary tissue and cultured neuronal and astroglial cells. The 27K BP expressed by BRL 3A cells (rBP-30) was found to also be expressed in conditioned media from newborn rat astrocytes and fetal neurons, but not in the medium from the neuroblastoma cell line, B104. Moreover, a polyclonal antibody, anti HEC1, specifically immunoprecipitated the BRL 3A BP from the same conditioned media, as well as from rat cerebrospinal and amniotic fluid and from conditioned medium of cells isolated from the neurointermediate lobe of adult rat pituitary. The same antibody also immunoprecipitated hBP-31 from human CSF. Northern blot analyses showed that rBP-30 mRNA was expressed in adult rat brain and pituitary, fetal brain and liver, and in fetal neurons and newborn astrocytes maintained in culture. We conclude that the BRL 3A BP (rBP-30) is the major insulin-like growth factor binding protein in the rat CNS and may be the rat analog of hBP-31, the predominant BP in human CSF.

The ovarian expression of the antigonadotropic insulin-like growth factor binding protein-2 is theca-interstitial cell-selective: evidence for hormonal regulation.

To begin the process of identification and charactization of rat ovarian insulin-like growth factor binding proteins, we have undertaken to explore the ovarian expression, cellular localization, and hormonal regulation of the insulin-like growth factor binding protein-2 (IGFBP-2) gene for which an antigonadotropic potential has recently been demonstrated. To this end, a solution hybridization/RNase protection assay was employed wherein total ovarian RNA (20 micrograms) from immature (21-23 days old) female rats was hybridized with a [32P]-labeled IGFBP-2 riboprobe. As in liver, a single protected fragment (550 bases long) corresponding to IGFBP-2 transcripts was identified in whole ovarian material. Cellular localization studies revealed the IGFBP-2 gene to be exclusively expressed in the theca-interstitial rather than the granulosa cell compartment. To confirm the cellular distribution of the IGFBP-2 protein, media conditioned by cultured granulosa or theca-interstitial cells were subjected to immunoprecipitation using two IGFBP-2-directed polyclonal antisera. Expectedly, both antibodies (but not non-immune rabbit serum) readily immunoprecipitated the 28 kDa rat IGFBP-2 species generated by hepatic BRL-3A cells. Similarly, both antibodies effectively immunoprecipitated an IGFBP the size of rat IGFBP-2 elaborated by theca-interstitial cells. In contrast, neither antibody immunoprecipitated the 28-29 kDa IGFBP species elaborated by granulosa cells otherwise readily apparent in conventional Western ligand blots. Hypophysectomy resulted in a 3-fold decrease (P less than 0.05) in the relative (densitometrically-quantified) abundance of ovarian IGFBP-2 transcripts, a diametrically opposed effect (P less than 0.05) being noted at the level of the liver. In contrast, treatment of immature hypophysectomized rats with a diethylstilbestrol-containing subcutaneous silastic implant for a total of 5 days resulted in a concordant 3-fold increase (P less than 0.05) in the relative abundance of IGFBP-2 transcripts in both ovary and liver when compared with untreated hypophysectomized controls. Taken together, these findings document rat ovarian IGFPB-2 gene expression to be theca-interstitial (rather than granulosa) cell-selective, and subject to upregulatory control by pituitary principle(s) and/or by estrogens. Although equally estrogen-dependent, hepatic IGFBP-2 gene expression proved constitutive in nature and subject to (diametrically opposed) inhibitory control by (potentially distinct) pituitary principle(s).

Elevated serum diiodotyrosine (DIT) in severe infections and sepsis: DIT, a possible new marker of leukocyte activity.

Ether link cleavage (ELC) of T4 yielding diiodotyrosine (DIT) has recently been shown in vitro to be the major pathway of T4 metabolism in phagocytosing leukocytes. To evaluate this pathway in vivo and the possible clinical relevance of DIT measurements in diseases with increased leukocyte activity, radioimmunological studies on serum levels of DIT and other thyroid parameters were performed in 125 critically ill patients classified into 3 groups with bacterial infections according to the severity of infection and 1 group without infections. While the pattern of iodothyronine and TSH levels typical for severe nonthyroidal disorders, i.e. decreased total T3 and elevated rT3, normal or decreased total T4 and TSH, and normal free T4, was found in all four groups of intensive care patients studied, elevated serum DIT was observed only in those patients whose clinical course was complicated by severe bacterial infections. Serial measurements revealed a close temporal connection between the infection phase and increased DIT levels. Median values and 16th to 84th percentile ranges (in parentheses) of serum DIT (normal range, 0.02-0.55 nmol/L) were as follows: sepsis, 1.38 (0.32-5.14); severe nonsystemic infections such as peritonitis and abscesses, 3.84 (0.24-17.2); moderate infections such as pneumonia and tracheobronchitis, 0.44 (0.18-1.16); and critical illness without infections, 0.14 (0.08-0.30) nmol/L. These elevations of circulating DIT could neither be correlated with changes in renal function nor attributed to drug effects. The results of the present study do not allow any definitive conclusions to be made about the mechanisms underlying the phenomenon of increased serum DIT levels in infections. Apart from this open question, DIT appears to be a relatively specific serum parameter for the presence and course of severe bacterial inflammations. Its measurement could provide useful clinical information, particularly for monitoring the time course of deep-seated infections.

Cloning and characterization of the gene encoding murine insulin-like growth factor-binding protein-2, mIGFBP-2.

We present a characterization of the single-copy gene, mIGFBP-2, encoding the murine insulin-like growth factor-binding protein-2 (mIGFBP-2). It consists of four exons with sizes of 470 +/- 2, 227, 141 and > 475 nucleotides (nt). The first intron spans 23 kb of genomic sequence, and the complete gene extends to more than 28 kb. Two kb of the 5'-flanking region were sequenced. This region has no TATA or CAAT boxes but is G+G-rich and contains several potential regulatory sequence motifs. A total of five GC boxes, which may serve as potential binding sites for a transcription factor, Sp1, are present immediately upstream of the transcription start point (tsp). By primer extension, we identified a single tsp at nt position -85 +/- 2. The murine IGFBP-2 locus was mapped to the proximal region of mouse chromosome 1, to a region of conserved synteny with human chromosome 2q. A comparison of the deduced amino acid sequences of mouse, rat and human IGFBP-2 reveals a high degree of homology between all three species.

Regulation of hepatic expression of IGF I and fetal IGF binding protein mRNA in streptozotocin-diabetic rats.

Hepatic mRNA levels of insulin-like growth factor I (IGF I) and of the fetal, nonglycosylated 32 kDa IGF-binding protein (BP) were analysed in diabetic, diabetic insulin- and IGF I-treated rats as well as in age-matched, healthy control animals. IGF ImRNA levels are reduced in diabetic rats and increased by insulin treatment. In contrast, the infusion of IGF I does not significantly upregulate IGF I mRNA levels. Fetal IGF BP mRNA expression is very low in healthy control animals, but high levels are found in diabetic rats. Insulin therapy lowers fetal IGF BP mRNA levels, whereas IGF I has no effect. We propose that insulin is a major regulator of the 32 kDa IGF BP levels in adult rats.

Differential regulation of insulin-like growth factor binding protein (IGFBP)-2 mRNA in liver and bone cells by insulin and retinoic acid in vitro.

Isolated cells produce insulin-like growth factors (IGFs) and their binding proteins (IGFBPs). Two distinct cell types were studied with regard to IGFBP-2 expression: (i) rat hepatocytes, which produce IGF I at a high rate and thus regulate its plasma concentration; and (ii) rat osteoblasts, which are targets of IGF I action. IGFBP-2 expression is low in hepatocytes prepared from normal adult rats and high in calvaria cells from newborn rats. Retinoic acid stimulates IGFBP-2 production by liver cells. Insulin suppresses both basal and retinoic acid-induced IGFBP-2 mRNA expression in hepatocytes and has no such effect on osteoblasts. Retinoic acid and insulin regulate IGFBP-2 expression in a tissue-specific manner.

Insulin-like growth factor (IGF)-I, -II and IGF binding protein-2 (IGFBP-2) in the plasma of children with Wilms' tumour.

Insulin-like growth factors (IGF)-I, -II and IGF binding protein-2 (IGFBP-2) have been measured in plasma of children with Wilms' tumour. The mean levels for total serum IGF-I and -II were not significantly altered in Wilms' tumour as compared with normal control plasma. However, the chromatographic profiles for IGF-I and -II in these groups were different with regard to the presence of IGF binding proteins and high molecular weight forms of IGFs; the high molecular weight form (9-15 kD) of IGF-II was significantly reduced in Wilms' tumour. Levels of IGFBP-2 were substantially elevated in serum from Wilms' tumour patients (1025 +/- 112 ng/ml compared with 416 +/- 44 ng/ml in controls), and inversely correlated with the levels of high molecular weight forms of IGF-II. We suggest that IGFBP-2 measurements might be of value as a marker for monitoring this type of tumour, either as an adjunct to diagnosis or surveillance of tumour growth during therapy.

Identification and characterization of the putative retinoblastoma control element of the rat insulin-like growth factor binding protein-2 gene.

The authors previously identified a silencer of the rat IGFBP-2 gene. Sequence examination of the silencer has revealed that it contains the target sequence for the pRb (retinoblastoma) tumour suppressor gene, referred to as the retinoblastoma control element (RCE) which is frequently found in the regulatory element of cellular oncogenes and growth factors. The presence of RCE suggests that the IGFBP-2 gene may be regulated by the pRb tumour suppressor gene. An in vitro gel retardation assay has shown that the putative RCEs from the IGFBP-2 gene are complexed with multiple nuclear factors from the rat liver BRL-3A cells. These DNA-protein complexes were not detected with the nuclear extracts from the cells that were growth arrested at the G1/S border of the cell cycle by high cell density. Using specific antibodies, Sp1 was shown to be one of the components for the multiple DNA-protein complex while pRb does not appear to be directly involved in the formation of the complex.

Influence of insulin-like growth factor-binding protein-2 on plasma clearance and transfer of insulin-like growth factors-I and -II from plasma into mammary-derived lymph and milk of goats.

Plasma clearance of insulin-like growth factors-I and -II (IGF-I and -II) and insulin-like growth factor-binding protein-2 (IGFBP-2) from lactating goats (n = 4) was determined following a single intravenous injection of the corresponding 125I-labelled human protein. Transfer of these proteins out of the vascular space was monitored by their subsequent appearance in mammary-derived lymph and milk. Clearance of 125I-IGFBP-2 from circulation was 0.37 +/- 0.06 ml/min/kg, which is markedly greater than that of 125I-IGF-I or -II (0.11 +/- 0.01 and 0.12 +/- 0.01 ml/min/kg respectively). This was also reflected in longer elimination half-lives for IGF-I (353 +/- 6 min) and -II (254 +/- 8 min) compared with IGFBP-2 (110 +/- 9 min). Three hours after injection of the 125I-labelled protein, the plasma:lymph ratio of trichloroacetic acid-precipitable radioactivity was 1.54 +/- 0.04, 3.3 +/- 0.6 and 4.1 +/- 0.4 for IGFBP-2, IGF-I and -II respectively. The form of 125I-IGFBP-2 in lymph was not different from that of plasma. Elevation of plasma concentrations of IGFBP-2 by its intravenous infusion significantly decreased plasma half-life of both IGF-I and -II (251 +/- 8 and 198 +/- 7 min respectively). Although the amount and rate of transfer of IGF into mammary-derived lymph was decreased slightly by IGFBP-2, concentrations eventually obtained were not different from control. However, secretion of IGFs into milk was significantly reduced by IGFBP-2, particularly in the case of IGF-I. These results are consistent with the ability of all three compounds to cross the vascular endothelium intact and of IGFBP-2 to decrease the uptake of IGF by mammary epithelium and subsequent secretion into milk. IGFBP-2 may well have acted to target plasma IGF towards non-mammary tissues, thus explaining the more rapid plasma clearance of IGFs in the presence of elevated IGFBP-2.

Circulating levels of IGFs and IGF binding proteins in human cord serum: relationships to intrauterine growth.

Cord sera were obtained from 44 term, human infants exhibiting various patterns of intrauterine growth and were assayed for IGF-1, IGF-2, and IGFBP-1, 2, and 3 by specific RIAs. Serum levels were correlated with birth weight (BW), ponderal index (PI), and placental weight (PW). Total IGF-1 levels correlated significantly with BW (r = 0.392), PW (r = 0.351), and PI (r = 0.481). By contrast, the correlation of IGF-2 with birth weight was not statistically significant (r = 0.264, P = 0.091). The association of IGF-2 with PI, however, was significant (r = 3.348, P = 0.024). IGFBP-3 exhibited significant correlations with BW, PI, and PW, similar to those seen with IGF-1. IGFBP-1 and IGFBP-2, however, were not significantly related to growth parameters. IGF-1 levels correlated strongly with IGFBP-3 levels (r = 0.646, P = 0.001). By contrast, IGF-1 correlated with the reciprocal of both IGFBP-1 and IGFBP-2. Based upon in vitro affinity constants, theoretical concentrations for each [IGF/IGFBP] complex, free IGFs, and free IGFBPs were calculated for each infant. Multiple regression analysis was performed including all 11 calculated variables and correlated with each growth parameter. This analysis revealed that an integrated expression of IGF activity exhibited stronger correlations with growth than each individual peptide species (BW, r = 0.681; PI, r = 0.660; PW, r = 0.658). These data further support roles for IGF related peptides (IGFRPs) in human fetal and placental growth and suggest regulatory/counterregulatory roles for the IGFBPs. It also supports the hypothesis that individual IGFRPs interact in a complex manner to define 'net IGF activity' in relation to fetal growth and/or metabolic status.