PubMed Health⌕ Search

Biomedical subjects

M Freemark

Publications and source records attributed to M Freemark.

At least 37 records · Page 2Linked to original sources
PubMed1993

Rat placental lactogen-I binds to the choroid plexus and hypothalamus of the pregnant rat.

Recent findings suggest that placental lactogen has a role in the regulation of hypothalamic function during pregnancy. To explore the mechanisms by which placental hormones may exert effects in the maternal central nervous system, we have examined the binding of rat placental lactogen-I (rPL-I) to brain slices from pregnant rats at mid- and late gestation. The binding of rPL-I to maternal rat brain was compared with that of human GH (hGH). Radiolabelled rPL-I bound specifically to ependymal cells of the choroid plexus in the lateral ventricles and in the roof of the third ventricle. The binding of 125I-labelled rPL-I was inhibited by unlabelled rPL-I, hGH or rat prolactin but not by rat GH, indicating that rPL-I and rat prolactin interact with a common binding site in maternal rat brain. Radiolabelled hGH bound to the choroid plexus and to ependymal cells lining the third ventricle in the region of the arcuate nucleus. In addition, hGH bound specifically to the ventromedial nuclei and to the medial preoptic area of the hypothalamus. The binding of radiolabelled hGH to all brain regions was inhibited by unlabelled rPL-I as well as hGH, indicating that rPL-I competes for lactogenic binding sites in the hypothalamus as well as the choroid plexus of the pregnant rat. These findings suggest potential mechanisms by which placental hormones may exert direct effects on the maternal central nervous system during pregnancy. The precise functions and roles of the PL-I binding sites in maternal choroid plexus and hypothalamus remain to be explored.

Animals↗
PubMed1992

Developmental regulation of insulin-like growth factor binding protein production: studies in fetal, postnatal, and pregnant sheep.

To assess the roles of developmental factors in the regulation of sheep IGFBP production at the cellular level, we characterized and compared the IGFBPs released by fetal, postnatal, and maternal sheep skin fibroblasts in culture with those in fetal, postnatal, and maternal sheep plasma. Sheep fibroblasts produced seven IGFBPs: a 36.5-41 kDa protein induced in vitro by IGF-I, likely representing oIGFBP-3; a 28.5 kDa protein that reacted with antisera to human IGFBP-2, likely representing oIGFBP-2; 25 and 27 kDa proteins induced in fetal fibroblasts by IGF-I; a 22 kDa protein that was inhibited by IGF-I, likely representing oIGFBP-4; and 21 and 23 kDa proteins labelled only by IGF-II, suggesting their similarities to IGFBP-6. The developmental pattern of IGFBP production by sheep fibroblasts in culture was similar in several respects to that observed in sheep plasma. For example, relative amounts of the 21, 22, and 28.5 kDa IGFBPs exceeded that of the 36.5-41 kDa protein in early fetal fibroblast conditioned media and in fetal plasma, while the relative concentrations of the 36.5-41 kDa protein increased markedly during the perinatal period. Sheep plasma differed, however, in two major respects from fibroblast conditioned media: First, fetal, and to a far lesser extent maternal, plasma contained a 200 kDa IGF-II-selective BP, likely to be the circulating form of the IGF-II receptor; and second, plasma, unlike conditioned media, contained a 26 kDa IGFBP, likely to be oIGFBP-1. The results of our studies suggest that the production and release of IGFBPs by isolated sheep fibroblasts is regulated by developmental factors operative under in vitro culture conditions. The differences in the relative levels of IGFBPs in conditioned media from fetal, postnatal, and maternal sheep fibroblasts resemble in several respects the differences in the relative concentrations of the various IGFBPs in fetal, postnatal, and maternal sheep plasma. Thus, sheep fibroblasts provide a useful though imperfect model system by which to examine the nutritional and hormonal regulation of sheep IGFBP production at various developmental stages.

Animals↗
PubMed1992

Binding of placental lactogen and growth hormone to fetal sheep fibroblasts.

Growth hormone (GH) regulates growth and development in the postnatal period but lacks somatotropic activity in the fetus. In contrast, the placental hormone placental lactogen (PL) stimulates amino acid transport, DNA synthesis, and somatomedin production in isolated fetal tissues, suggesting that PL may function as a "fetal GH." To elucidate the mechanisms by which PL exerts GH-like effects in fetal tissues, we examined the binding of PL, GH, and prolactin to cultured skin fibroblasts obtained from midgestational fetal lambs. Ovine fetal fibroblasts bound radiolabeled ovine PL (oPL) specifically and with high affinity (EC50 0.20 nM). In competitive displacement assays using 125I-oPL as the radioligand, the potency of unlabeled oPL was eight to 12 times greater than that of ovine GH and congruent to 1000 times greater than that of ovine prolactin. Covalent cross-linking of 125I-oPL (22 kD) to ovine fetal fibroblasts revealed a specific hormone-receptor complex with an apparent M(r) of 130,000, suggesting that the high affinity oPL binding site has a molecular mass of approximately 108 kD. The specific bindings of radiolabeled ovine GH (0.6% per 250 micrograms protein) and ovine prolactin (0.04% per 250 micrograms protein) were only 1/15 and 1/230 that of radiolabeled oPL (9.1% per 250 micrograms protein), and no specific cross-linking of 125I-ovine GH or 125I-ovine prolactin to ovine fetal fibroblasts was detected. These findings demonstrate preferential binding of PL by isolated fetal sheep fibroblasts in culture, providing a cellular mechanism whereby PL may exert growth-promoting effects in the fetus.

Animals↗
PubMed1992

The placental lactogen receptor in maternal and fetal sheep liver: regulation by glucose and role in the pathogenesis of fasting during pregnancy.

To clarify the roles of glucose and insulin in the regulation of the PL receptor in fetal and maternal sheep liver, we administered iv glucose to pregnant ewes during a 72-h fast. The binding of ovine PL (oPL) to hepatic membranes from glucose-infused ewes and their fetuses was compared with the binding of oPL to tissues of fasted, saline-infused sheep and sheep fed normally ad libitum. Fasting of pregnant ewes caused a 58-70% reduction in the number of PL receptors in fetal and maternal liver. Intravenous administration of glucose during fasting increased the number of PL receptors in fetal liver by 137.4%. In contrast, glucose administration during fasting had no effect on the number of PL receptors in maternal liver. The number of PL binding sites in fetal liver correlated positively with fetal weight (r = 0.59) and length (r = 0.54) and with fetal plasma glucose (r = 0.69) and insulin (r = 0.55) concentrations. In contrast, PL binding was inversely related to fetal plasma oPL concentrations (r = -0.70). These findings suggested that glucose, insulin, and/or oPL may regulate PL binding in the ovine fetus. To determine whether glucose or insulin exert direct effects on the PL receptor in ovine fetal tissues, we examined the binding of radiolabeled oPL to ovine fetal hepatocytes and fibroblasts in culture. The specific binding of oPL to fetal hepatocytes was low and variable (1.0 +/- 0.5%) and it was not possible to assess reliably the effects of glucose or insulin supplementation. The specific binding of oPL to fetal fibroblasts (5.4 +/- 0.6%/mg) was unaffected by variations in media glucose concentrations (5.5-16.5 mM) or by pretreatment with insulin (10-1000 ng/ml). The results of these studies demonstrate that glucose and other nutritional factors regulate the expression of the PL receptor in fetal and maternal sheep liver. Alterations in PL binding play roles in the metabolic adaptation of the mother and fetus to nutritional deprivation and stress.

Animals↗
PubMed1992

Nutritional regulation of insulin-like growth factor-binding protein gene expression in the ovine fetus and pregnant ewe.

The factors controlling the synthesis and degradation of the insulin-like growth factor-binding proteins (IGFBPs) during pregnancy are poorly understood. To clarify the roles of nutritional factors in the regulation of fetal and maternal IGFBP production, we examined the effects of fasting, refeeding, and glucose administration on plasma IGFBP concentrations and hepatic IGFBP mRNA levels in fetal lambs and pregnant ewes (n = 24). Maternal fasting for 3 days in late gestation stimulated a 50-100% increase in maternal plasma BP-1 concentrations (P < 0.05) and a 2- to 3-fold increase in fetal plasma BP-1 (P < 0.05), as determined by densitometric analysis of Western ligand blots. Fasting also stimulated a 40-70% increase in maternal plasma BP-2 concentrations (P < 0.05), but had no significant effect on fetal plasma BP-2 levels. Levels of hepatic BP-1 mRNA in the fetus and pregnant ewe during fasting paralleled plasma BP-1 levels, suggesting that fasting modulates fetal and maternal plasma BP concentrations at least in part through effects on hepatic gene expression. The effects of fasting on both mRNA and plasma levels of BP-1 and BP-2 were reversed by 3 days of refeeding and were prevented by glucose infusion during fasting. When ewes were made hyperglycemic by the infusion of hypertonic glucose, plasma BP-1 and BP-2 concentrations varied inversely with blood glucose concentrations. In addition, hyperglycemia reduced maternal liver BP-1 and BP-2 mRNA levels and fetal BP-1 mRNA levels by 50-65%. Direct administration of hypertonic glucose to the fetus decreased fetal plasma BP-1 levels acutely and reduced fetal BP-1 mRNA levels by 57%, but had no effect on fetal plasma BP-2 or fetal hepatic BP-2 mRNA levels. These findings indicate that glucose and other nutritional factors regulate gene expression and plasma levels of BP-1 and BP-2 in the pregnant ewe and BP-1 in the fetal lamb. The changes in expression of these IGFBPs during fasting and hyperglycemia may play roles in adaptation of the pregnant mother and fetus to metabolic stress.

Animal Nutritional Physiological Phenomena↗
PubMed1992

Evidence for a novel insulin-like growth factor (IGF)-dependent protease regulating IGF-binding protein-4 in dermal fibroblasts.

The mechanisms by which insulin-like growth factors (IGFs) reduce IGF-binding protein-4 (IGFBP-4) levels in cellular conditioned media are poorly understood. The effect of IGFs on IGFBP-4 levels in fibroblast conditioned media is not mediated via the type 1 or type 2 cellular IGF receptors, and the IGFs exert little or no effects on IGFBP-4 messenger RNA levels in human adult fibroblasts or in rat neuroblastoma cells. To determine whether the effects of IGFs on IGFBP-4 might be exerted through alterations in IGFBP-4 degradation, we incubated cell-free, fibroblast-conditioned media from either sheep or human dermal fibroblasts with or without IGF-I, IGF-II (each 1 microgram/ml), or insulin (10 micrograms/ml) for 72 h at 37 C. Samples were then analyzed by Western ligand blot using radiolabeled IGFs and by immunoblotting using a polyclonal antisera to human IGFBP-4. In the absence of IGFs, no apparent changes in the basal concentrations of the various IGFBPs were observed. In contrast, incubation of media with IGFs caused a 70-80% reduction in levels of both sheep and human IGFBP-4, whereas incubation with insulin was without effect. Similarly, incubation of cell-free conditioned media containing recombinant human IGFBP-4 with IGF-I caused a reduction in detectable levels of the 28K protein. The decrease in IGFBP-4 levels was accompanied by the appearance of an immunoreactive approximate 17-20K fragment that did not bind radiolabeled IGFs by ligand blot. The IGF-dependent decrease in IGFBP-4 was prevented by coincubation of the media with serine protease inhibitors, EDTA, or 1,10-phenanthrolene, suggesting that IGFs may activate an IGFBP-4 specific metallo-serine protease present in fibroblast conditioned media. Alternatively, binding of IGF-I or -II to IGFBP-4 may enhance the susceptibility of IGFBP-4 to proteolytic degradation. The demonstration that IGF-I and IGF-II can promote directly the proteolytic degradation of IGFBP-4 into fragments that do not bind IGFs provides a novel mechanism by which the IGFs may increase their own availability and/or activity in biological fluids.

Animals↗
PubMed1990

Placental lactogen receptors in maternal sheep liver: effects of fasting and refeeding.

In a recent study we demonstrated that fasting of the pregnant ewe reduces the number of placental lactogen (PL) receptors in fetal sheep liver. In the present study we examined the effects of a 72-h fast on the number and affinity of PL receptors in maternal sheep liver. Fasting caused a 57% reduction in the number of hepatic ovine PL receptors; this effect was reversed by refeeding. There were no changes in the affinity of the PL receptor, the subunit structure of the receptor, or the extent of occupancy of the receptor by endogenous circulating maternal hormones. The number of hepatic PL receptors was correlated positively with the maternal plasma concentrations of glucose and insulin, suggesting that these factors may regulate PL binding to maternal tissues during pregnancy. In addition, PL receptor number was correlated positively with maternal plasma insulin-like growth factor I (IGF-I) concentrations, suggesting that a reduction in hepatic ovine PL binding may contribute to the reduction in maternal IGF-I during fasting. Fasting produced a 72% reduction in the number of ovine growth hormone receptors in maternal liver and an 83% increase in hepatic insulin binding. These findings indicate that fasting of the pregnant ewe reduces the number of PL receptors in maternal and fetal liver. The reduction in PL binding may contribute to maternal and fetal hyposomatomedinemia and may play a role in the pathogenesis of the intrauterine growth retardation that accompanies maternal nutritional deprivation.

Animals↗
PubMed1989

Purification of a distinct placental lactogen receptor, a new member of the growth hormone/prolactin receptor family.

Recent findings from this laboratory suggest that the biological actions of placental lactogen (PL) in mammalian fetal tissues are mediated through binding of the hormone to a distinct and unique PL receptor. We have now purified this receptor from fetal and maternal sheep liver, characterized its binding to PL, growth hormone (GH), and prolactin (PRL), and determined its molecular weight by SDS-PAGE and by affinity cross-linking techniques. Soluble extracts containing specific, high-affinity (Kd 0.5 nM) PL binding activity were prepared by incubating ovine fetal and maternal liver microsomes with 1% Triton X-100. The detergent solubilized PL receptor was purified two- to threefold by ion-exchange chromatography and an additional twofold by gel exclusion chromatography on Sepharose 6B. The PL receptor was then purified 75,000- to 125,000-fold by affinity chromatography using a column of ovine PL (oPL) coupled to Affi-Gel 10. The molecular weight of the oPL receptor as determined by SDS-PAGE and by cross-linking techniques was 44,000 +/- 2,000 (range 40,000-48,000). The purified receptor bound 125I-oPL specifically and with high affinity (Kd 0.5 nM) but did not bind either radiolabeled ovine GH or ovine PRL. In addition, in competition studies using 125I-oPL as the radioligand, the purified PL receptor bound unlabeled oPL with a potency 30-50 times greater than that of ovine GH and 500-1,000 times greater than that of ovine PRL. These findings demonstrate the presence of a specific PL receptor in fetal and maternal sheep liver. The PL receptor, together with the GH and PRL receptors, constitute a family of distinct but related hormone receptors that differ in their relative affinities for PL, GH, and PRL. Changes in the expression of the three receptors may mediate changes in the hormonal control of growth during the transition from fetal to postnatal life.

Animals↗
PubMed1989

Nutritional regulation of the placental lactogen receptor in fetal liver: implications for fetal metabolism and growth.

We have recently identified and purified from fetal liver a distinct receptor that mediates the effects of placental lactogen (PL) on amino acid transport, glycogen synthesis, and somatomedin production in fetal tissues. At present, the factors that regulate the number and affinity of PL receptors in the fetus are unknown. Since maternal nutrition plays a critical role in fetal metabolism and growth, we have examined the role of nutrition in the regulation of the PL receptor in fetal lambs. Pregnant ewes at 123-126 days gestation were fed ad libitum (FED), fasted for 3 days (FASTED), or fasted for 3 days and then refed for an additional 3 days (REFED). The ewes were then killed, and the binding of [125I]ovine (o) PL to hepatic microsomes from the fetal lambs was examined. Maternal fasting caused a 60-75% reduction in the specific binding of oPL to fetal liver; the effect of fasting was reversed in part by refeeding [specific binding per mg protein: FED, 11.8 +/- 2.2% (n = 8); FASTED, 2.8 +/- 0.4% (n = 7); REFED, 7.2 +/- 2.6% (n = 3)]. The decrease in oPL binding resulted from an 80% reduction in the number of fetal oPL-binding sites (Scatchard analysis); there were no changes in the affinity of the oPL receptor (Kd, 0.6 nM), the subunit structure of the receptor, or the degree of occupancy of the receptor in vivo by endogenous fetal hormones. The specific bindings of GH (0.6%), PRL (0.3%), and insulin (35%) to fetal liver were not affected by maternal fasting, indicating that caloric restriction exerted a specific effect on oPL binding in the fetus. The number of fetal oPL-binding sites was positively correlated with the fetal liver glycogen content (r = 0.69; P less than 0.01) and the fetal plasma concentrations of glucose (r = 0.68; P less than 0.01) and insulin-like growth factor-I (r = 0.74; P less than 0.001), suggesting a role for the PL receptor in the regulation of fetal carbohydrate metabolism and growth. The number of fetal PL receptors was inversely correlated with the fetal plasma oPL concentration (r = 0.47; P less than 0.05). These studies demonstrate that fasting of the pregnant ewe reduces the number of PL receptors in ovine fetal liver. The reduction in fetal hepatic PL receptors may contribute to the mobilization and depletion of fetal liver glycogen stores and may play a role in the pathogenesis of the fetal growth retardation that accompanies maternal caloric deprivation.

Animals↗
PubMed1988

Differential solubilization of placental lactogen (PL)- and growth hormone-binding sites: further evidence for a unique PL receptor in fetal and maternal liver.

Previous studies from this laboratory provided evidence for the existence of a specific placental lactogen (PL) receptor in tissues of fetal lambs and pregnant sheep. The PL receptor is structurally and functionally distinct from somatotropic (GH) and lactogenic (PRL) receptors, and there are conspicuous differences in the expression of the three receptors during ontogeny. The results of the present study indicate striking differences in the solubilization of PL- and GH-binding sites in maternal and fetal sheep liver. Radiolabeled ovine PL (oPL) bound specifically and with high affinity (Kd, 0.97 nM) to soluble detergent extracts of ovine fetal liver, but there was no specific binding of radiolabeled ovine GH (oGH) or oPRL to soluble extracts or insoluble fractions of fetal liver. When liver microsomes of pregnant sheep were extracted with Triton X-100, 80% of the [125I]oPL-binding sites were recovered in the soluble fraction, but 76% of the [125I]oGH binding sites were recovered in the insoluble pellet. Soluble extracts of maternal liver had high affinity for oPL (Kd, 1.45 nM), but low affinity for oGH (Kd 33 nM) and oPRL (Kd, 1-2 microM). On the other hand, Triton-insoluble fractions of maternal liver had high affinity for oGH (Kd, 0.95 nM) as well as oPL (Kd, 0.91 nM), but low affinity for oPRL (Kd, 1-2 microM). The subunit structure of the [125I]oPL-binding site in soluble fractions of fetal and maternal liver (mol wt, 38-47K) was distinct from that of the [125I]oGH-binding site in Triton-insoluble fractions of maternal liver (mol wt, 54/118K). These findings indicate that treatment of microsomal fractions of fetal and maternal sheep liver with Triton X-100 solubilizes the oPL receptor but not the oGH receptor. The differential solubilization of PL- and GH-binding sites may facilitate purification of the two distinct receptors and clarification of their respective roles in the regulation of fetal and postnatal growth.

Animals↗
PubMed1988

Placental lactogen and growth hormone receptors in human fetal tissues: relationship to fetal plasma human placental lactogen concentrations and fetal growth.

The specific binding of human placental lactogen (hPL) and human GH (hGH) to particulate cell membranes from human fetal liver and skeletal muscle at 12-19 weeks gestation was examined. Fetal liver and muscle specifically bound [125I]hPL. This binding was inhibited by increasing concentrations of unlabeled hPL (half-maximal concentrations, 2.2 and 3.4 nmol/L, respectively). Scatchard analysis of the hepatic membrane binding revealed curvilinear plots with higher (Kd, 2.2 nmol/L) and lower (Kd, 24 nmol/L) affinity sites, while binding to muscle involved a single receptor class of Kd 5.6 nmol/L. The binding capacities for the two hepatic sites correlated positively with fetal body weight. [125I]hGH specifically bound to liver, but not muscle, with higher (Kd, 1.6 nmol/L) and lower (Kd, 8.6 nmol/L) affinity sites. [125I]PRL bound to hepatic membranes, but was preferentially displaced by hPL or hGH. Between 4 and 500 micrograms/L (mean, 82 micrograms/L, 3.8 nmol/L) hPL were present in fetal plasma. The findings identify distinct hPL receptors in human fetal liver and skeletal muscle and a hepatic hGH receptor in midgestation.

Embryonic and Fetal Development↗
PubMed1987

Role of placental lactogen and prolactin in human pregnancy.

In summary, studies from our and other laboratories strongly suggest that placental lactogen has direct effects on fetal growth and metabolism as well as on maternal metabolism. Prolactin may be important in the regulation of water and ion transport across the amnion, the production of surfactant by the fetal lung, and the immune response during pregnancy. A summary of the postulated effects of placental lactogen on maternal and fetal physiology is depicted in Figure 9 and a summary of the postulated effects of prolactin during pregnancy is shown in Table 6. Undoubtedly, future studies of the physiology of placental lactogen and prolactin will uncover new functions for these hormones during gestation.

Female↗
PubMed1987

Epidermal growth factor (EGF)-like transforming growth factor (TGF) activity and EGF receptors in ovine fetal tissues: possible role for TGF in ovine fetal development.

To determine whether epidermal growth factor (EGF) or EGF-like transforming growth factors (TGFs) are present in ovine fetal tissues, we have tested acid-ethanol extracts of ovine fetal kidney for the ability to induce anchorage-independent growth of normal rat kidney fibroblasts in soft agar and to compete with 125I-mouse EGF for binding to receptors in sheep liver. The fetal kidney extract (20-1000 micrograms protein/ml) stimulated a dose-dependent increase in the number of soft agar colonies of normal rat kidney cells. Approximately 70% of these colonies measured greater than or equal to 3100 microM2. The fetal kidney extract was dissolved in 1 M acetic acid and chromatographed on Bio-Gel P 10. Two peaks of TGF-like activity, with approximate MW 14.5K (peak 1) and 9K (peak 2), eluted from the column. Half-maximal effects of pooled peaks 1 and 2 on colony formation were achieved using 100 and 20 micrograms protein/ml, respectively. Peaks 1 and 2 also competed with 125I-mouse EGF for binding to EGF receptors in ovine fetal liver but had no activity in a homologous mouse EGF radioimmunoassay sensitive to 15 pg mouse EGF. Neither TGF activity nor EGF receptor binding activity was detected in Bio-Gel fractions co-eluting with 125I-mouse EGF. Specific 125I-EGF binding sites in fetal liver were detected as early as midgestation, and the number of EGF binding sites increased markedly in late gestation, exceeding the number of EGF binding sites in the livers of pregnant ewes. These findings demonstrate the presence of TGF-like activity in ovine fetal kidney and high affinity EGF receptors in ovine fetal liver.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1987

A unique placental lactogen receptor: implications for fetal growth.

To determine whether there are structural differences between the binding sites for placental lactogen (PL) and GH, we have compared the molecular weights of complexes formed by the covalent cross-linking of [125I]ovine (o) PL and [125I]oGH to hepatic membranes from fetal and pregnant sheep in mid- and late gestation and from postnatal nonpregnant sheep at 3 days to 7 months of age. Specific [125I]oPL binding sites in fetal liver were detected as early as midgestation, and cross-linking of [125I]oPL to fetal hepatic membranes yielded a major radiographic band with a mol wt of 60 +/- 5 K (mean +/- SD). Unlabeled oPL at low concentrations (0.9-9 nM) specifically competed with [125I]oPL for binding to the 60 K complex. In contrast, oGH and oPRL competed for binding to the 60 K complex only at much higher concentrations (greater than or equal to 90 nM). In addition, no specific cross-linking of [125I]oGH or [125I]oPRL to fetal hepatic membranes was observed. These findings suggest the presence of a distinct and unique PL binding site in ovine fetal liver. Since the mol wt of oPL is 22 K, the estimated mol wt of the oPL receptor protein is 38 +/- 5 K. During the first week after birth, there was a striking increase in the number of [125I]oGH binding sites. Cross-linking of [125I]oGH to postnatal liver yielded radiographic bands with apparent mol wts of 75 K and 140 K. The relative potencies of oPL, oGH, and oPRL in competing for binding to the 75 K and 140 K complexes were similar to the relative potencies of these hormones in competing for [125I]oGH binding sites in postnatal liver, suggesting that the 75 K and 140 K bands represent subunits of the oGH receptor bound covalently to [125I]oGH. Cross-linking of [125I]oPL to pregnant and postnatal nonpregnant liver yielded three radiographic bands with mol wts of 60 K, 75 K, and 140 K. The intensities of all three bands were reduced by low concentrations (0.9-9 nM) of oPL. Higher concentrations of oGH abolished the 75 K and 140 K bands but reduced the intensity of the 60 K band by only 20-30%. oPRL had minimal effect on band intensities. These observations suggest the presence of two functionally and structurally distinct receptors in pregnant liver: the oPL receptor, which has high affinity for oPL and low affinity for oGH and oPRL.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1986

Placental lactogen and GH receptors in sheep liver: striking differences in ontogeny and function.

To determine whether changes in the relative biological potencies of ovine placental lactogen (oPL) and ovine growth hormone (oGH) during development derive from ontogenetic changes in the binding of these hormones to hepatic receptors, we have compared the binding of 125I-oPL and 125I-oGH to hepatic membranes from fetal lambs and pregnant sheep at mid- and late gestation and from postnatal sheep at 1 day to 7 mo of age. Specific high-affinity 125I-oPL binding sites in ovine fetal liver were detected as early as day 70 of gestation (term = 145 days), and the number of fetal 125I-oPL binding sites increased progressively throughout the latter half of gestation, reaching a maximum (11.2 fmol/mg protein) at 3-7 days before parturition. The potency of oPL (Kd 0.27 nM) in competing for 125I-oPL binding sites was 90 and 1,300 times greater than that of oGH and ovine prolactin, respectively. Although the number of fetal 125I-oPL binding sites increased throughout pregnancy, there was little or no specific binding of 125I-oGH noted in the fetus. Treatment of fetal liver membranes with 4 M MgCl2 did not enhance the subsequent specific binding of 125I-oGH, suggesting that the low specific binding of oGH did not result from occupation of hepatic receptors by endogenous circulating oPL or oGH. In contrast, MgCL2 treatment markedly increased the apparent number of fetal 125I-oPL binding sites, suggesting that oPL receptors in fetal liver are partly saturated in vivo by oPL.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1986

The glycogenic effects of placental lactogen and growth hormone in ovine fetal liver are mediated through binding to specific fetal ovine placental lactogen receptors.

To examine the relative roles of placental lactogen (PL) and GH in fetal metabolism, we have examined the effects of ovine PL (oPL), ovine GH (oGH), and ovine PRL (oPRL) on glycogen metabolism in cultured ovine fetal hepatocytes and have examined the binding of these hormones to hepatic membranes from fetal and neonatal lambs. In ovine fetal hepatocytes, oPL (150 ng/ml-20 micrograms/ml) stimulated dose-dependent increases in [14C]glucose incorporation into glycogen (18-167%) and total cellular glycogen content (10-69%). oGH and oPRL also stimulated glycogen synthesis in fetal hepatocytes, but the potencies of these hormones were only 12% and 4% that of oPL. The dose-response curves of the three hormones were parallel, and their maximal effects were identical, suggesting a common mechanism of action. In hepatic membranes from fetal lambs, the maximal specific binding of [125I]oPL was 26.3% while the maximal specific binding of [125I]oGH was only 0.9-1.5%. The binding of [125I]oPL was saturable and reversible and varied with incubation time and temperature. Unlabeled oPL (1 ng/ml-5 micrograms/ml) caused a dose-dependent inhibition of the binding of [125I]oPL to fetal hepatic membranes, with half-maximal displacement of [125I]oPL by 5-7 ng unlabeled oPL/ml. oGH and oPRL caused parallel displacement of [125I]oPL, but with potencies only 2% and 0.1% that of oPL. Scatchard analysis of oPL dose-response curves indicated that the hormone bound to a single class of receptors with a dissociation constant of 1.1 X 10(-10) M. The maximal specific binding of [125I]oGH to hepatic membranes of neonatal lambs (20.1%) greatly exceeded the binding of oGH to fetal hepatic membranes. In addition, the potency of oGH in competing for [125I]oPL binding sites in neonatal liver greatly exceeded the potency of oGH in competing for [125I]oPL binding sites in fetal liver. Although the biological effects of both oPL and oGH in postnatal subprimate tissues may be mediated through binding to nonprimate GH receptors, the results of these studies suggest that the glycogenic effects of oPL in ovine fetal liver are mediated through binding to specific fetal oPL receptors. The relatively weak biological effects of oGH and oPRL in ovine fetal liver appear to be mediated through the binding of the hormones to fetal oPL receptors. The presence of specific, high affinity PL receptors in ovine fetal tissues provides a mechanism whereby oPL may function as a GH in the ovine fetus.

Animals↗
PubMed1986

Epidermal growth factor stimulates glycogen synthesis in fetal rat hepatocytes: comparison with the glycogenic effects of insulin-like growth factor I and insulin.

The effects of epidermal growth factor (EGF) on glycogen metabolism and the binding of [125I]iodo-EGF to receptors in fetal rat hepatocytes have been examined. The actions of EGF have been compared with those of insulin-like growth factor I (IGF-I) and insulin. EGF (0.1-45 nM) stimulated dose-dependent increases in [14C]glucose incorporation into glycogen (8.8-31.1%, P less than 0.01) and total cellular glycogen content (5.6-21.4%, P less than 0.05). The concentration of EGF causing half-maximal stimulation of glycogen synthesis was 2 ng/ml, and maximal stimulation occurred at 1 h of incubation. EGF had no effect on the uptake of the nonmetabolizable monosaccharide [14C]O-methyl-D-glucose, suggesting that the glycogenic effect of EGF was not mediated through stimulation of glucose transport. Although IGF-I (1-100 nM) and insulin (14 nM to 10 microM) also stimulated glycogen synthesis in fetal liver, the maximal effects of these hormones occurred at 2 h incubation, and the dose-response curves of IGF-I and insulin were not parallel to that of EGF. In addition, the maximal glycogenic effect of EGF was only 40% that of insulin or IGF-I, and the effects of EGF and insulin on [14C]glucose incorporation were additive. These findings suggest that EGF stimulates glycogen synthesis through a mechanism distinct from that of IGF-I or insulin. The binding of [125I]iodo-EGF to fetal hepatocytes was specific, saturable, and time- and temperature-dependent. Maximal specific binding occurred at 1 h of incubation at 37 C or at 24 h of incubation at 4 C. Unlabeled EGF (0.05-250 ng/ml) caused a dose-dependent inhibition of the binding of [125I]iodo-EGF to fetal hepatocytes, with half-maximal displacement of [125I]iodo-EGF by 1.7 ng unlabeled EGF/ml. The specific binding of [125I] iodo-EGF was not inhibited by high concentrations of insulin or IGF-I, suggesting that the differences in the mechanisms by which EGF, insulin, and IGF-I stimulate glycogenesis may be explained in part by differences in the binding of these hormones to fetal liver receptors. In addition to having mitogenic effects in fetal tissue, EGF or other EGF-like growth factors may have acute effects on fetal hepatic intermediary metabolism and may contribute to the accumulation of liver glycogen in the mammalian fetus during late gestation.

3-O-Methylglucose↗