Hepatic IGF-I mRNA Levels Research Articles

Modulating effect of elevated water hardness on growth performance, ammonia dynamics and ion-regulatory capacity in channel catfish (Ictalurus punctatus) following chronic challenge with high environmental ammonia and salinity stress

The aim of this study was to assess whether rearing fish at increased water hardness could assist channel catfish (Ictalurus punctatus) juveniles to efficiently cope with later (sub)lethal ammonia exposure or salinity stress. Fish were subjected to 100, 494 (10% of 96 h-LC50) or 1250 mg/L (25% of 96 h-LC50) CaCO3 water hardness levels for two months. Afterwards, fish were challenged with high environmental ammonia (HEA; 12.2 mg/L total ammonia ~25% of 96 h-LC50 value) or salinity stress (SST;10 ppt) separately for 21 days at their respective water hardness levels. Results show that after two months, weight gain (%), specific growth rate and feed conversion in 1250 mg/L CaCO3 group were markedly worse than 100 and 494 mg/L CaCO3 groups. This finding was accompanied with a down-regulation of hepatic IGF-I and growth hormone receptors (GHR) mRNA levels. During SST, growth was still significantly worse for those at 1250 mg/L CaCO3 (1250SST). However, exposure to HEA inhibited growth for 100HEA and 494HEA relative to 1250HEA. Ammonia excretion rate (Jamm) remained similar at different hardness levels; however, this was strongly inhibited at 100HEA, 494HEA and 100SST. This appeared to align to some degree with high water hardness having a protective effect on the gill histomorphology. In fact, fish at 1250HEA were able to increase Jamm efficiently, which was associated with upregulated branchial expression of ammonia transporters (Rhesus glycoproteins ‘Rhcg’) and Na+/H+ exchanger, as well as augmented gill H+-ATPase activity. These responses prevented a build-up of excess ammonia in plasma. In contrast, both 1250 mg/L CaCO3 and 1250HEA groups displayed inhibited branchial Na+/K+-ATPase activity indicating this unlikely has a major role in ammonia excretion. Gill Ca2+-ATPase activities reduced at the highest hardness (1250 mg/L CaCO3) and 1250HEA groups, likely to prevent hypercalcemia in plasma. No differences among hardness levels were noted for ion-regulation or Ca2+- homeostasis under SST. Overall, findings suggest that although elevated water hardness up to 1250 mg/L CaCO3 adversely affects the growth performance of channel catfish, it can ameliorate the inhibitory growth effects of HEA. This was partly attributed to 1250 mg/L CaCO3 protecting the gills as well as mitigating HEA-induced ammonia excretory and ion-regulatory disruption. However, elevated hardness may not be as effective in alleviating salinity stress in catfish.

Evidences for involvement of growth hormone and insulin-like growth factor in ovarian development of starry flounder (Platichthys stellatus).

Although gonadotrophins are major regulators of ovarian function in teleosts and other vertebrates, accumulating evidence indicates that the growth hormone (GH)-insulin-like growth factor (IGF) axis also plays an important role in fish reproduction. As a first step to understand the physiological role of the GH-IGF system in the ovarian development of starry flounder (Platichthys stellatus), the expression profiles of GH and IGF messenger RNAs (mRNAs) and plasma GH, IGF-I, estradiol-17β (E2), and testosterone (T) levels during the ovarian development were investigated. The developmental stages of ovaries were divided into five stages (II, III, IV, V, and VI) by histological analysis. The hepatosomatic index (HSI) and gonadosomatic index (GSI) values increased and peaked at stage IV and stage V, respectively, and then declined at stage VI. Pituitary GH mRNA levels decreased sharply at stage III and raised to top level at stage VI. The hepatic IGF-I mRNA levels ascended to maximum value at stage V and then declined significantly at stage VI. However, the hepatic IGF-II mRNA levels remained stable and increased significantly at stage VI. In contrast, the ovarian IGF-I mRNA levels increased gradually and peaked at stage VI. The ovarian IGF-II mRNA levels were initially stable and increased significantly at stage V until the top level at stage VI. Consistent with the pituitary GH mRNA levels, plasma GH levels reduced sharply at stage III and remained depressed until stage V and then raised remarkably at stage VI. Plasma IGF-I level peaked at stage V and then declined to initial level. Plasma E2 level peaked at stage IV and then dramatically descended to the basal level. Plasma T level peaked at stage V and then declined significantly back to the basal level. Based on statistical analysis, significant positive correlations between hepatic IGF-I mRNA and GSI, ovarian IGF-II mRNA and hepatic IGF-II mRNA, ovarian IGF-I mRNA and ovarian IGF-II mRNA, and plasma IGF-I and plasma T were observed, respectively. These results suggest that the GH-IGF system may be involved in the ovarian development of starry flounder; GH and IGFs appear to play distinct roles in the regulation of the ovarian development in paracrine/autocrine manners. These findings extend our knowledge of the roles of the GH-IGF axis on reproduction regulation in fish.

Effects of dietary arginine levels and carbohydrate-to-lipid ratios on mRNA expression of growth-related hormones in largemouth bass, Micropterus salmoides

Utilizing the tissue samples and growth data collected from our two preceding researches in largemouth bass (LMB), we have investigated effects of dietary arginine (Arg) levels and carbohydrate-to-lipid (CHO/LIP) ratios on the GH, IGF-I and insulin expression in related tissues to find possible relationships between the nutrient intake, growth performance and transcript level. Hepatic IGF-I and pituitary GH mRNA levels were significantly up-regulated by lower dietary Arg levels from 1.94% to 3.01% and by higher levels from 2.76% to 3.01%, respectively, while Brockmann body (BB)-containing tissue insulin mRNA expression was not affected. Dietary CHO/LIP ratios ranging from 0.32 to 5.17 (w/w) affected pituitary GH, liver IGF-I and BB-containing tissue insulin mRNA expression in a ratio-specific pattern. The lower ratios from 0.32 to 2.36 significantly up-regulated GH and insulin transcript levels, but significantly down-regulated IGF-I transcript levels; the higher ratios did no longer exert any further effects on them. Meanwhile, two strong positive correlations (r=0.892, r=0.885) between hepatic IGF-I transcript levels and specific growth rates of tested fish were observed with varying dietary Arg levels and CHO/LIP ratios, respectively. These findings indicate that in LMB dietary Arg levels and CHO/LIP ratios regulate differentially the endocrine system of GH, IGF-I and insulin at transcription level; this system, in turn, plays a fundamental role in the regulation of the nutrient metabolism and somatic growth; and that hepatic IGF-I mRNA abundance should be a more reliable index to assess growth and nutritional fitness than the others, at least in LMB.

Dietary Carbohydrate Source Alters Gene Expression Profile of Intestinal Epithelium in Mice

High-sucrose consumption is associated with increased risk of human colon cancer. Our previous research indicated that high-sucrose diets (vs. cornstarch) promote intestinal epithelial cell proliferation and tumorigenesis as well as increase serum glucose and hepatic IGF-I mRNA levels in APC Min mice. To examine the role of functional pathways, in particular of IGF-I signaling, in sucrose-induced intestinal epithelial cell proliferation and tumorigenesis, we examined the effects of dietary carbohydrate source (sucrose vs. cornstarch) on gene expression in the intestinal epithelium using cDNA microarray and quantitative RT-PCR analysis. Dietary carbohydrate source significantly ( P < 0.05) altered mRNA expression of 109 known genes in the small intestinal epithelium, including many involved in metabolic pathways. Consumption of high-sucrose diets altered expression levels of genes involved in cell adhesion, cell cycle control, and transduction signaling, consistent with increased risk of intestinal tumorigenesis. High-sucrose intake also affected expression of genes involved in IGF-I signaling, including upregulating IGF-II and downregulating IGFBP3, which supports our hypothesis that IGF-I signaling could play a role in intestinal epithelial cell proliferation and tumorigenesis promoted by high-sucrose consumption.

Molecular characterization and gene expression analysis of insulin-like growth factors I and II in the redbanded seabream, Pagrus auriga: transcriptional regulation by growth hormone

Insulin-like growth factors (IGFs) I and II (IGF-I and IGF-II) play important roles in fish growth and development. The present study was aimed at isolating cDNAs encoding both IGF-I and IGF-II in the redbanded seabream ( Pagrus auriga), and at measuring relative gene expression levels in different organs and during larval development. A fragment of 1321 nucleotides coding for IGF-I was cloned from liver using 3′ and 5′ RACE techniques. It included an open reading frame of 558 nucleotides, encoding a 185-amino acid preproIGF-I. With respect to IGF-II, a fragment of 1544 nucleotides was cloned as well. The open reading frame spanned 648 nucleotides, rendering a 215-amino acid preproIGF-II. The deduced mature 67-amino acid IGF-I and 70-amino acid IGF-II exhibited high sequence identities with their corresponding fish counterparts, ranging between 88.6-100% and 79.1–98.5%, respectively. Real-time PCR showed the highest IGF-I transcripts in liver (∼ 200-fold higher than head-kidney). In contrast, the highest IGF-II mRNAs were detected in gills and heart (∼ 16-fold higher than head-kidney). In addition, both IGFs exhibited different gene expression patterns during larval development suggesting that their expression is developmentally regulated. IGF-I reached the highest expression levels at 18 days after hatching (11.6-fold higher than 1 day after hatching), whereas IGF-II expression did not change significantly. Both hepatic IGF-I and IGF-II mRNA levels increased sharply (3.1- and 19-fold higher than control, respectively) 3 h after injection of porcine growth hormone, but remained unchanged from 6 to 24 h after treatment. Our results are discussed in relation to those previously reported for other bony fish.

Overexpression of IGF‐I transgene rescued growth retardation of weaver mutant mice

IGF-I is an important growth factor for postnatal growth in mammals. To evaluate the relative contribution of IGF-1 in postnatal growth during pathological states, we used weaver mutant mouse. We found that, compared with their control littermates, the body weights of both male and female weaver (wv) mice were significantly decreased soon after birth and remained low later in life, although male wv mice showed a trend to catch-up to normal growth. Organ weights of wv mice also showed a proportional decrease. To determine if the growth retardation in wv mice is related to IGF-I deficiency, we measured circulating IGF-I levels by radioimmunoassay and hepatic IGF-I mRNA levels by Northern blot. Out data showed a parallel decrease of IGF-I and body weights in female and D21 male wv mice. After male wv mice entered puberty, their IGF-I levels caught-up to that of their wildtype littermates as well as their body weight. These data suggested that IGF-I deficiency may be the cause of postnatal growth retardation in wv mutant mice. After crossbred wv mice with IGF-I transgenic mice, wv mice carrying the IGF-I transgene restored their body weights to those of their wildtype littermates. Interestingly, among other organs, only brain, spleen and female liver showed significant weight increases. In summary, we demonstrated that the decrease in circulating IGF-I contributed to postnatal growth retardation in wv mutant mice, which was confirmed by the nearly normal growth pattern of wv mice in the presence of IGF-I transgene. Overall, our results support the notion that IGF-I plays an important role in postnatal growth in normal as well as neurodegenerative states.

Insulin-like growth factor-I cDNA cloning, gene expression and potential use as a growth rate indicator in Nile tilapia, Oreochromis niloticus

IGF-I is a mitogenic polypeptide that is an important regulator of growth in fish. The potential of IGF-I mRNA abundance as a rapid growth indicator in the Nile tilapia, Oreochromis niloticus, was evaluated. Hepatic IGF-I cDNA was isolated and partially cloned. The partial sequence having 539 bases encodes for the signal peptide, mature protein and a portion of the E domain. The deduced 68 amino acid sequence for mature IGF-I showed 84–90% and 77–79% sequence identity with fish and mammalian counterparts, respectively. The deduced amino acid sequence for domains B and A was most conserved (93–97%) relative to other fishes. A sensitive TaqMan real time qRT-PCR assay for O. niloticus was developed based on the mature IGF-I peptide for measures of hepatic IGF-I mRNA levels. Hepatic IGF-I mRNA levels were found to be significantly correlated with growth rate of fish reared under different feeding regimes and temperature conditions. Higher feed consumption and water temperature produced faster-growing fish and increased hepatic IGF-I mRNA expression. These findings suggest that hepatic-derived IGF-I plays a key role in controlling growth in O. niloticus and indicates that IGF-I mRNA quantification could prove useful for the rapid assessment of growth rate in this species.

Intact Insulin and Insulin-Like Growth Factor-I Receptor Signaling Is Required for Growth Hormone Effects on Skeletal Muscle Growth and Functionin Vivo

GH and IGF-I are potent regulators of muscle growth and function. Although IGF-I is known to mediate many of the effects of GH, it is not yet clear whether all effects of GH are completely dependent on the IGF-I system. To evaluate the biological effects of the GH/IGF-I axis on muscle growth, we administrated recombinant human GH to mice, which lack IGF-I function specifically in skeletal muscle, due to the overexpression of a dominant-negative IGF-I receptor in this tissue (MKR mice). GH treatment significantly increased the levels of hepatic IGF-I mRNA and serum IGF-I levels in both wild-type (WT) and MKR mice. These GH-induced effects were paralleled by increases in body weight and in the weights of most GH-responsive organs in both groups of mice. Interestingly, unlike WT mice, GH treatment had no effect on skeletal muscle weight in MKR mice. GH treatment failed to reverse the impaired muscle function in MKR mice. Furthermore, MKR mice exhibited no effects of GH on the cross-sectional area of myofibers and the proliferation of satellite cells. Taken together, these data suggest that the in vivo effects of GH on muscle mass and strength are primarily mediated by activation of the IGF-I receptor.

Diurnal Variation in Growth Hormone Receptor Messenger Ribonucleic Acid in Liver and Skeletal Muscle of lit/+ and lit/lit Mice

The present study investigated the diurnal variation in GH receptor (GHR) mRNA in liver and skeletal muscle of 3-month-old GH-deficient and -sufficient mice using quantitative real-time RT-PCR. lit/lit (GH deficient) or lit/+ (GH sufficient) mice were fed ad libitum and lights were on between 0600 and 2000. Tissues were collected at 0800-1000, 1200-1400 and 2000-2200. Hepatic GHR mRNA levels of lit/+ mice at 0800-1000 were significantly lower than those at 1200-1400 and 2000-2200. There was no significant variation in hepatic GHR mRNA of lit/lit mice. In skeletal muscle, GHR mRNA levels of both lit/+ and lit/lit mice at 1200-1400 were significantly higher than those at 0800-1000 and 2000-2200. There was also a diurnal change in hepatic IGF-I mRNA levels of lit/+ but not of lit/lit mice; the levels were lowest at 0800-1000 in lit/+ mice. On the other hand, there was no variation in IGF-I mRNA levels in skeletal muscle. These results suggest that 1) there is a diurnal variation in GHR expression in liver and skeletal muscle and the pattern of the variation is tissue specific; 2) GH deficiency blunted the diurnal variation in GHR mRNA in liver but not that in skeletal muscle; 3) IGF-I mRNA expression in liver is more closely related to GHR mRNA expression than that in skeletal muscle.

IL-6-overexpression brings about growth impairment potentially through a GH receptor defect.

This paper is concerned with growth retardation associated with overproduction of interleukin-6 (IL-6). As a model, we used MUP/hIL-6 transgenic mice in which human IL-6 cDNA is overexpressed under the control of a MUP gene enhancer/promoter. The growth-retardation of MUP/hIL-6 transgenic mice was paralleled by reduced serum levels of IGF-I. As shown, hepatic IGF-I mRNA levels were reduced in the transgenic mice. MUP/hIL-6 transgenic mice are in a state of growth hormone (GH)-resistance, since their serum GH levels are either normal or elevated. To identify possible steps in GH signaling which might be perturbed in the transgenic mice, we examined the synthesis of GH receptor (GHR) mRNA. We noted a twofold reduction of hepatic GHR mRNA in the transgenic mice. We therefore conclude that overexpression of IL-6 brings about growth impairment in part through a GH receptor defect.

PCR-cloning and gene expression studies in common carp ( Cyprinuscarpio) insulin-like growth factor-II

Insulin-like growth factor-II (IGF-II) is a member of a growth factor family related to fetal growth in mammals but its physiological role has not been clearly identified in fish. In teleosts, the basic mechanism of the growth hormone (GH)–IGF axis is known to be operative but in a different manner. For instance, IGF-I exhibits GH dependence whereas for IGF-II, its GH dependence varies in different fish species. In this study, we used polymerase chain reaction (PCR) to obtain a common carp IGF-II (ccIGF-II) cDNA fragment and methods of rapid amplification of cDNA ends (RACEs) to obtain a full-length ccIGF-II sequence. The ccIGF-II encodes for a predicted amino acid sequence showing identities of 70.6%, 68.7%, 63.4% and 35% in comparison with salmon, barramundi, tilapia and human IGF-II, respectively. The nucleotide identity between the open reading frame (ORF) of the ccIGF-II and ccIGF-I cDNA sequence is only 36.2%. Distribution of ccIGF-II mRNA levels in common carp tissues was also studied; ccIGF-II expressed in hepatopancreas, heart, and many other tissues in adult carps are similar to the levels of ccIGF-I except in gills and testis. ccIGF-II levels were significantly higher than that of ccIGF-I in most juvenile tissues except in hepatopancreas, where ccIGF-I was higher (threefold) than that of ccIGF-II. The levels of ccIGF-I were also higher than ccIGF-II in carp larvae, from pre-hatched stage to day 30 post-hatching. Injection of porcine GH (pGH) increased the IGF-I and IGF-II mRNA levels in the hepatopancreas and brain of juvenile carps. However, hepatic IGF-I mRNA levels were induced more than IGF-II by pGH, whereas ccIGF-II levels gave a higher response than IGF-I in the brain in response to GH induction.

Insulin-like growth factor mRNA in Barramundi (Lates calcarifer): alternative splicing and nonresponsiveness to growth hormone.

Barramundi (Lates calcarifer) is a teleost of the superorder Acanthopterygii. Barramundi IGF-I cDNA was cloned and the distribution of alternative transcripts in various barramundi tissues was investigated using rt-PCR and RPA. It was demonstrated that in barramundi tissues, IGF-I mRNAs were represented by two transcripts corresponding to the reported salmonid Ea-2 and Ea-4. The acute effect of GH on hepatic IGF-I mRNA levels was investigated. Seven hours after intraperitonal administration of either 6 micrograms of recombinant bream GH/g body weight or saline, no significant increase in the levels of either of the two transcripts could be observed. The presence of GH receptors in the barramundi liver was demonstrated in binding assays using recombinant bream GH and liver membrane preparations. An analysis of a barramundi IGF-I genomic sequence encompassing the three exons that encode the E domain suggested that the pattern of splice site utilization is determined by the degree of homology of the splicing signals to the consensus splice site sequences.

Nutritional Regulation of Circulating Insulin-like Growth Factors (IGFs) and Their Binding Proteins in the Ovine Fetus.

Insulin-like growth factors (IGFs) and their binding proteins (IGFBPs) are important for fetal and postnatal development, but the regulation of circulating IGFs and IGFBPs has not been as thoroughly investigated in the maternal/fetal unit as in the adult animal where nutrition status plays a regulatory role. We used the chronically-catheterized, late-gestation ovine model and compared circulating IGFs and IGFBPs levels, and hepatic IGF-I mRNA levels. Following a five-day maternal fast, both IGF-I and IGF-II levels were decreased in the maternal and fetal circulation (P < 0.05), accompanied by a decrease in fetal hepatic IGF-I mRNA levels, but the IGFBP2 level was increased and the IGFBP3 level was decreased in maternal circulation, whereas the IGFBP1 level was increased in fetal circulation. In both fed and fasting states, the infusion of glucose (150% of baseline) did not alter IGFs or IGFBPs in either maternal or fetal circulation. To understand the regulation of the endogenous IGF system, rhIGF-I was infused (6.7 nmol/kg fetus/h) into the fetal circulation. While maternal IGFs or IGFBPs remained unchanged, IGF-I infusion into fetal circulation resulted in an increase in IGF-I, a decrease in IGF-II, and an overall increase in the IGFBPs (P < 0.05). Taken together, circulating IGFs and IGFBPs in the ovine fetus are more sensitive to prolonged nutrient deficit than to a brief glucose increase. The nutrition status therefore regulates the IGF system in maternal and fetal circulation which, in turn, may regulate the nutrient utilization for fetal growth.

Decreased GH-IGF-I Axis in Obese Rats Induced by High Fat Diet

Growth hormone (GH) and IGF-I are considered to be essential to adipocyte differentiation and may play a key role in fat mass accumulation in vivo. The aim of this study is to determine the effect of high lipid content diet (HL) on serum GH and tissue IGF-I mRNA levels in rats. Male Wistar rats were administered a diet containing 29% energy as lipid for 18 weeks. Significant body weight increase was observed in HL-treated rats at 28 weeks of age, comparing with normal control (CO) rats. Significantly increased fat accumlation was also shown in epididymal and subcutaneous adipose tissues in HL rats. Serum GH levels in HL rats were significantly lower than those in CO rats (HL=16.5±2.9 (SEM)ng/ml vs CO=31.7±6.0, p<0.01). Total RNA was extracted and applied to northern blot to hybridize with a rat IGF-I cDNA probe. IGF-I mRNAs were expressed at 7.5 kb with several minor bands in liver and white adipose tissue. Hepatic IGF-I mRNA/β-actin mRNA ratio in HL rats was significantly decreased (HL=0.74±0.07 vs CO=1.66±0.59, p<0.01) in accordance with decreased serum GH levels. The epididymal fat IGF-I mRNA/β-actin mRNA ratio in HL was also significantly lower than those in CO (HL=0.56±0.13 vs CO=0.81±0.14, p<0.02), whereas there was no significant difference in subcutaneous fat IGF-I mRNA/β-actin mRNA ratio between HL and CO (HL=1.61±0.31 vs CO=1.68±0.42, NS). IGF-I mRNA levels in both liver and epididymal fat in HL rats decreased proportionately to serum GH levels, suggesting that hepatic and epididymal fat IGF-I mRNA levels are mainly regulated by GH. In an obese state, increased visceral fat accumulation may be promoted by decreased serum GH levels or vice versa.

Hypoxia-induced changes in insulin-like growth factors and their binding proteins in pregnant rats.

Pregnancy is associated with important changes in the insulin-like growth factor (IGF)-insulin-like growth factor binding protein (IGFBP) axis, but the importance of these growth factors for fetal growth is not well understood. We have recently established a maternal hypoxia model that results in significant intrauterine growth retardation in the fetus, and characterized the IGF-IGFBP axis in growth-retarded fetuses. To determine if maternal IGFs and their binding proteins are similarly regulated by hypoxia, we examined their expression in 6 hypoxic dams (13% oxygen, days 14-21 of gestation) and 6 control dams (21% oxygen). There was no significant difference in the food intake between the groups. The mean body weight of hypoxic dams, however, was 20% less than that of controls. Of all the organs, the lungs were most affected by hypoxia, weighing 17% more in the hypoxic dams than in the control dams; placental weight was reduced by 10% in the hypoxic dams. Liver and brain weights were not changed significantly by hypoxia. The mean concentration ofimmunoreactive IGF-I was 123 +/- 11 ng/ml in the hypoxic dams and 130 +/- 18 ng/ml in the control dams (nonsignificant). Similarly, there was no significant difference in hepatic IGF-I mRNA levels determined by solution hybridization nuclease-protection assay. An increase in IGFBP-1, IGFBP-2 and IGFBP-4 concentrations, however, could be observed by Western ligand blotting of the sera of hypoxic dams, compared to control dams. As assessed by Northern blot analysis, there was a 2.8-fold increase in IGFBP-1 mRNA expression in the livers of hypoxic dams compared to controls. Hepatic IGFBP-4 expression was also slightly increased (1.25-fold) in the hypoxic dams. No difference in hepatic IGFBP-2 or IGFBP-3 mRNA was found. Our results show parallel patterns in fetal and maternal IGF and IGFBP responses to hypoxia. This suggests that hypoxia may inhibit fetal growth by both directly affecting the fetus and via inhibition of placental growth.

Reduction in circulating IGF-I and hepatic IGF-I mRNA levels after caecal ligation and puncture are associated with differential regulation of hepatic IGF-binding protein-1, -2 and -3 mRNA levels.

Sepsis is characterized by a severe shift in metabolism, characterized by low IGF-I levels. We have studied the influence of caecal ligation and puncture (CLP) on the levels of circulating IGF-I and hepatic IGF-I and IGF-binding protein (IGFBP)-1, -2 and -3 mRNA in adult male Wistar rats (n = 12) and compared it with sham-operated rats (n = 6). In order to exclude anorexia-induced changes we also studied animals pair-fed to both groups. IGF-I levels were measured by RIA. Steady-state hepatic IGF-I, IGFBP-1, IGFBP-2 and IGFBP-3 mRNA levels were measured by Northern blot analysis using specific rat cDNA probes. Food intake averaged 13.0 +/- 2.0 g/day in the sham-operated rats fed ad libitum during the study period, with a sharp decline in food intake in the CLP animals (2.3 +/- 1.3 g/day). After CLP, there was a significant reduction in circulating IGF-I levels (467.2 +/- 50.9 micrograms/l) compared with sham-operated animals (924.0 +/- 75.3 micrograms/l; P = 0.04) or those pair-fed to the CLP rats (612.5 +/- 52.9 micrograms/l; P = 0.04). Total hepatic IGF-I mRNA levels were significantly reduced (2.57 +/- 0.05 densitometric units (DU) after CLP compared with the sham-operated group (2.71 +/- 0.04); P = 0.04), or their pair-fed controls (2.75 +/- 0.08 DU; P < 0.05). Hepatic IGFBP-3 mRNA levels were lower after CLP (0.37 +/- 0.04 DU) than in the sham-operated animals (0.66 +/- 0.09 DU; P = 0.04) or their pair-fed controls (0.61 +/- 0.05 DU; P = 0.04). On the other hand, hepatic IGFBP-2 mRNA levels were increased after CLP (0.91 +/- 0.11 DU) compared with sham-operated animals (0.28 +/- 0.06 DU; P = 0.01) or with their pair-fed controls (0.22 +/- 0.02; P = 0.01), as were hepatic IGFBP-1 mRNA levels (CLP animals 0.95 +/- 0.11 DU; sham-operated 0.30 +/- 0.04 DU, P = 0.01; pair-fed 0.30 +/- 0.02 DU, P = 0.01). No significant difference between sham-operated animals and their pair-fed controls was observed in circulating IGF-I levels (888.0 +/- 109.3 micrograms/l; P = not significant (N.S.)), hepatic mRNA levels for IGF-I (2.72 +/- 0.06 DU; P = N.S), IGFBP-3 (0.71 +/- 0.07 DU; P = N.S.), IGFBP-2 (0.25 +/- 0.07 DU; P = N.S.) or IGFBP-1 (0.27 +/- 0.06 DU; P = N.S.). In summary, after CLP there was a reduction in both circulating and hepatic IGF-I mRNA levels associated with a specific and differential regulation of hepatic IGFBP-1, -2 and -3 mRNA levels. Although we cannot eliminate a possible effect of surgical stress combined with malnutrition, our results suggest that these changes are a specific effect of sepsis rather than simply a result of surgical stress or poor nutrition alone.

Influence of nutrition on hepatic IGF-I mRNA levels and plasma concentrations of IGF-I and IGF-II in meat-type chickens.

We have examined the influence of nutrition on plasma IGF-I, IGF-II and IGF-binding protein (IGFBP) levels and on hepatic IGF-I gene expression in young meat-type chickens. Plasma IGF concentrations were measured by using RIA with recombinant chicken IGFs as standards. In chickens fed the control diet containing 200 g/kg dietary protein ad libitum for 7 days, plasma IGF-I concentrations increased significantly from those found in the initial control group. Food restriction for either 4 or 7 days decreased plasma IGF-I by 30% from the initial control. When chickens were refed ad libitum for 3 days after 4 days of restricted feeding, plasma IGF-I levels recovered to those of the control birds fed ad libitum. In chickens eating a low protein diet (100 g/kg protein), the plasma IGF-I tended to be lowered but the decrease was not significant. Although the intensity of IGF-I and beta-actin mRNA bands protected in the RNase protection assay was changed by nutrition, no statistical effect of nutrition on the ratio of IGF-I to beta-actin was observed. The nutritional treatments had no effect on plasma IGF-II concentrations. Western ligand blot and chromatographic analyses were used to investigate the influence of nutrition on IGFBP profiles. Both IGF-I and IGF-II ligands in the Western ligand blot revealed the most intense binding at 30 kDa for plasma obtained from chickens with restricted food intake. The 30 kDa band also appeared at a lower intensity in the group fed a low protein diet but not in any other groups. These observations were confirmed by neutral gel chromatography. The chicken IGF-II ligand revealed an intensely labelled band corresponding to 75 kDa and this was not affected by nutrition. IGF-I and IGFBP concentrations in the plasma of young broiler chickens were influenced by nutritional state but IGF-II concentrations were not. The lack of a response in circulating IGF-II levels may have been due to the presence of high concentrations of a 75 kDa specific binding protein which did not respond to nutrition in this experiment.

Differential expression and hormonal regulation of alternatively spliced IGF-I mRNA transcripts in salmon

Salmon have been shown to express alternatively spliced IGF-I mRNA transcripts coding for four different IGF-I prohormones. These transcripts, now designated Ea-1, Ea-2, Ea-3 and Ea-4, differ in size due to the inclusion of additional sequences in the E domain-coding region of the molecule. In this study, the tissue distribution and hormonal regulation of expression of alternatively spliced IGF-I mRNA transcripts were investigated in coho salmon. IGF-I mRNAs were detected by solution hybridization/RNase protection assay in all tissues examined. GH treatment significantly increased hepatic IGF-I mRNA content. Hepatic IGF-I mRNA levels were not influenced by prolactin or somatolactin. Heart, fat, brain, kidney, spleen and ovary IGF-I mRNA levels were not affected by GH, prolactin or somatolactin. Ea-1, Ea-3 and Ea-4 mRNA transcripts were detectable in the liver, and Ea-1 and Ea-3 levels increased dramatically in response to GH treatment, whereas the amount of Ea-4 mRNA was unchanged. Most non-hepatic tissues expressed only the Ea-4 transcript, and expression was not influenced by GH, prolactin or somatolactin. Ea-1 and Ea-3 transcripts were visible in gill samples from fish treated with GH. The ovaries of juvenile fish expressed Ea-1, Ea-2 and Ea-4. The amounts of these transcripts were not changed by gonadotrophin treatment. During smoltification of juvenile coho salmon, liver and gill IGF-I mRNA levels increased with increasing plasma GH and thyroxine concentrations. Muscle, brain and ovary IGF-I mRNA levels were unchanged during this period. These data suggest that the liver is a major site of IGF-I production in response to GH. Heart, fat, brain, kidney, spleen and ovary did not show increased IGF-I mRNA levels in response to GH treatment. GH and prolactin had inconsistent effects on muscle IGF-I mRNA levels. Somatolactin and a gonadotrophin preparation did not stimulate IGF-I expression in tissues of juvenile fish. Differences in tissue GH responsiveness can be partially explained by the expression of alternatively spliced IGF-I mRNAs. Of the four hepatic IGF-I mRNA transcripts, Ea-1 and Ea-3 are GH-responsive, while Ea-2 and Ea-4 are not. Most non-hepatic tissues express only the Ea-4 transcript, and IGF-I mRNA levels do not increase after GH treatment. The increased IGF-I mRNA levels observed in gill tissue during smoltification suggest that other factors, in addition to GH, may regulate IGF-I expression.(ABSTRACT TRUNCATED AT 400 WORDS)

The effect of ovarian function on insulin-like growth factor I plasma levels and hepatic IGF-I mRNA levels in diabetic rats treated with insulin

When insulin was administered to streptozotocin-induced diabetic female rats, the percentage of glycohemolobin, growth rate, ovulatory cycle, uterus to body weight ratio, and insulin-like growth factor (IGF-I) level returned to near normal. In untreated diabetic rats there were no normla estrous cycles, and hepatic IGF-I mRNA (8.94 ± 1.02 O.D. units per μg total RNA) levels were significantly lower than the control or insulin-treated groups in proestrus (16.47 ± 0.91 and 17.15 ± 1.84, respectively). Insulin therapy restored the hypothalamic-pituitary-ovarian axis with the reinstitution of normal estrous cycles. Plasma IGF-I levels were highest in non-diabetic proestrous animals (277 ± 36.9 ng/ml), significantly higher than IGF-I levels in insulin-treated diabetic rats in diestrus (174 ± 23.1 ng/ml), non-diabetic diestrous rats (165 ± 18.4 ng/ml) and untreated diabetic rats (135 ± 19.7 ng/ml). Plasma IGF-I levels were elevated in insulin-treated diabetic rats in proestrus (221 ± 78.3 ng/ml), however this was not significantly different from any other group. The increases observed in plasma IGF-I and hepatic IGF-I mRNA after insulin theraphy correlate with the normalization of sex hormone secretion. Though this study does not prove a causal relationship between restoration of ovarian function and normalization of circulating IGF-I levels, a relationship has been established, as evidenced by higher levels of IGF-I in both the control and insulin-treated diabetic proestrous groups when compared to the diestrous groups.

Regulation of insulin-like growth factor-I and growth hormone receptor gene expression by diabetes and nutritional state in rat tissues

Insulin-like growth factor-I (IGF-I) mRNA and GH receptor mRNA levels were analysed in different tissues from rats made diabetic with streptozotocin, fasted rats and rats fed with a protein-reduced diet. Diabetes decreased IGF-I mRNA levels in liver, heart, diaphragm, kidney and aorta, but not in brain. GH receptor mRNA levels were decreased in heart and diaphragm, but not in liver and kidney. Fasting decreased IGF-I mRNA in all tissues studied except brain, and decreased GH receptor mRNA in liver, heart and diaphragm, but not in kidney. A protein-reduced diet decreased hepatic IGF-I mRNA levels but did not significantly affect other tissues, while GH receptor mRNA levels were reduced in liver and diaphragm. In conclusion, both diabetes and limited nutrition affected IGF-I and GH receptor mRNA in different tissues, but the two mRNAs were not co-ordinately regulated in all tissues studied. While reduced GH receptor gene expression may thus be responsible for decreased IGF-I gene expression in some states and tissues, additional regulatory mechanisms may be of importance.