补偿生长对异育银鲫IGF-1、IGFBP-1水平及IGF-1、IGF-1RmRNA表达的影响

该实验分析饥饿和恢复投喂对异育银鲫血液IGF-1和IGFBP-1水平和肝脏IGF-1、白肌IGF-1RmRNA表达量的影响。结果显示:饥饿期(14d)血液中IGF-1和IGFBP-1水平逐渐下降,在饥饿第14天均出现显著性降低(P<0.05);恢复投喂后第1天IGF-1迅速恢复到对照组水平,而IGFBP-1水平仍显著低于对照组(P<0.05),随后逐渐升高,直至于恢复投喂第14天后显著高于对照组水平(P<0.05);饥饿期肝脏IGF-1mRNA表达量呈下降趋势,但与对照组无显著性差异(P>0.05);恢复投喂初期(第1、3天),IGF-1mRNA表达量仍继续下降(P<0.05),对营养条件的变化反应滞后,至第7天,表达水平恢复到对照组水平。白肌IGF-1RmRNA表达水平在饥饿第3天出现显著性下降(P<0.05),继续饥饿其水平出现补偿性升高;恢复投喂后第14天IGF-1RmRNA表达量显著高于对照组水平(P<0.05)。该结果揭示恢复投喂期高水平的IGFBP-1含量和IGF-1RmRNA表达量可能通过提高IGF-1的促生长作用参与异育银鲫的补偿生长调节。     

The metabolic effects of prolonged starvation and refeeding in sturgeon and rainbow trout

The present study examines the particular metabolic strategies of the sturgeon Acipenser naccarii in facing a period of prolonged starvation (72 days) and subsequent refeeding (60 days) compared to the trout Oncorhynchus mykiss response under similar conditions. Plasma metabolites, endogenous reserves, and the activity of intermediate enzymes in liver and white muscle were evaluated. This study shows the mobilization of tissue reserves during a starvation period in both species with an associated enzymatic response. The sturgeon displayed an early increase in hepatic glycolysis during starvation. The trout preferentially used lactate for gluconeogenesis in liver and white muscle. The sturgeon had higher lipid-degradation capacity and greater synthesis of hepatic ketone bodies than the trout, although this latter species also showed strong synthesis of ketone bodies during starvation. During refeeding, the metabolic activity present before starvation was recovered in both fish, with a reestablishment of tissue reserves, plasmatic parameters (glucemia and cholesterol), and enzymatic activities in the liver and muscle. A compensatory effect in enzymes regarding lipids, ketone bodies, and oxidative metabolism was displayed in the liver of both species. There are metabolic differences between sturgeon and trout that support the contention that the sturgeon has common characteristics with elasmobranchs and teleosts.     

Effect of starvation and re‐feeding on growth performance and content of plasma lipids,glucose and insulin in cultured juvenile Persian sturgeon (Acipenser persicus Borodin, 1897)

The effect of starvation and subsequent re‐feeding to satiation on compensatory growth performance, insulin and blood serum values were investigated in juvenile Persian sturgeon (Acipencer persicus) with an average weight 108.04 ± 0.28 g (mean ± SEM) and in the same rearing condition over an 8‐week period. Sturgeons were allocated to one of five feeding treatments: controls (C, continuous feeding), W1 (1 week starvation), W2 (2 weeks starvation), W3 (3 weeks starvation) and W4 (4 weeks starvation), followed by a single 4 weeks of re‐feeding to satiation. Changes in growth performance and blood serum indices were examined at the end of weeks 4 and 8. Body weight, specific growth rate (SGR), condition factor (CF) and weight gain were determined to have significantly decreased during starvation. Fish starved for 1 week reached the same weight as the control fish after re‐feeding for 4 weeks, indicating that complete compensatory growth occurred. Although the specific growth rate in W2, W3 and W4 fish was greater than that in the control fish after re‐feeding, W2, W3 and W4 fish did not reach the same body weight as control fish at the end of re‐feeding period, and showed partial compensation only. Blood plasma, glucose and insulin concentrations did not change significantly during starvation and re‐feeding (P > 0.05). This suggests that sturgeon are able to maintain glycaemia during starvation, probably due to their non‐carbohydrate dietary source. Plasma total lipid and triglyceride levels increased in starvation treatments, whereas the increases were significant only in W3 treatment (P < 0.05). After a 4‐week re‐feeding period, their levels decreased in comparison to the starvation periods. Increases in plasma total lipid and triglyceride levels appear to be due to their roles as preferred nutrients for mobilization in Persian sturgeon and the magnitude and duration of compensatory growth depended on the length of food deprivation.     

Energy reserves during food deprivation and compensatory growth in juvenile roach: the importance of season and temperature

The effect of 21 days of starvation, followed by a period of compensatory growth during refeeding, was studied in juvenile roach Rutilus rutilus during winter and summer, at 4, 20 and 27° C acclimation temperature and at a constant photoperiod (12L : 12D). Although light conditions were the same during summer and winter experiments and fish were acclimated to the same temperatures, there were significant differences in a range of variables between summer and winter. Generally winter fish were better prepared to face starvation than summer fish, especially when acclimated at a realistic cold season water temperature of 4° C. In winter, the cold acclimated fish had a two to three‐fold larger relative liver size with an approximately double fractional lipid content, in comparison to summer animals at the same temperature. Their white muscle protein and glycogen concentration, but not their lipid content, were significantly higher. Season, independent of photoperiod or reproductive cycle, was therefore an important factor that determined the physiological status of the animal, and should generally be taken into account when fish are acclimated to different temperature regimes. There were no significant differences between seasons with respect to growth. Juvenile roach showed compensatory growth at all three acclimation temperatures with maximal rates of compensatory growth at 27° C. The replenishment of body energy stores, which were utilized during the starvation period, was responsible for the observed mass gain at 4° C. The contribution of the different energy resources (protein, glycogen and lipid) was dependent on acclimation temperature. In 20 and 27° C acclimated roach, the energetic needs during food deprivation were met by metabolizing white muscle energy stores. While the concentration of white muscle glycogen had decreased after the fasting period, the concentrations of white muscle lipid and protein remained more or less constant. The mobilization of protein and fat was revealed by the reduced size of the muscle after fasting, which was reflected in a decrease in condition factor. At 20° C, liver lipids and glycogen were mobilized, which caused a decrease both in the relative liver size and in the concentration of these substrates. Liver size was also decreased after fasting in the 4° C acclimated fish, but the substrate concentrations remained stable. This experimental group additionally utilized white muscle glycogen during food deprivation. Almost all measured variables were back at the control level within 7 days of refeeding.     

Porcine insulin-like growth factor-I (pIGF-I): complementary deoxyribonucleic acid cloning and uterine expression of messenger ribonucleic acid encoding evolutionarily conserved IGF-I peptides

In order to facilitate studies of insulin-like growth factor-I (IGF-I) expression during the pregnancy-associated development of uterus and mammary gland in the pig model, we have isolated several cDNA clones corresponding to porcine IGF-I (pIGF-I) mRNA. Sequence analysis of two cDNA fragments (sigf. 2 and sigf. 3) revealed an open reading frame encoding in order a putative 25 amino acid (aa) hydrophobic leader peptide, the mature (processed) 70 aa pIGF-I peptide and a 35 aa carboxy-terminal extension (E) peptide. The deduced aa sequence of the pIGF-I peptide is identical to human and bovine IGF-I but differs from that of rat and mouse at three and four residues, respectively. The sequences of the amino- and carboxy-terminal IGF extension peptides are also highly conserved among these species. Northern analysis using sigf. 3 as a probe revealed multiple IGF-I mRNAs (including species of 8000, 2300, and 1200 nucleotides in length) in uteri of pregnant pigs. Highest levels of the uterine IGF-I mRNAs were found at early pregnancy, when increased levels of immunoreactive tissue IGF-I were also observed. Mammary levels of IGF-I mRNAs and protein were considerably lower than that observed for uterus at the same time period. Thus, uterine production of IGF-I appears to be especially significant during early pregnancy in the pig when uterine growth, elevated IGF-I in uterine fluids, and rapid embryonic development are observed.     

Real-time polymerase chain reaction, in situ hybridization and immunohistochemical localization of insulin-like growth factor-I and myostatin during development of Dicentrarchus labrax (Pisces: Osteichthyes)

The distribution of insulin-like growth factor-I (IGF-I) and myostatin (MSTN) was investigated in sea bass (Dicentrarchus labrax) by real-time polymerase chain reaction (PCR), in situ hybridization (ISH) and immunohistochemistry. Real-time PCR indicated that IGF-I mRNA increased from the second day post-hatching and that this trend became significant from day 4. ISH confirmed a strong IGF-I mRNA expression from the first week post-hatching, with the most abundant expression being detected in the liver of larvae and adults. Real-time PCR also showed that the level of MSTN mRNA increased significantly from day 25. The expression of MSTN mRNA was higher in muscle and almost absent in other anatomical regions in both larvae and adults. Interestingly, the lateral muscle showed a quantitative differential expression of IGF-I and MSTN mRNAs in red and white muscle, depending on the developmental stage examined. IGF-I immunoreactivity was detected in developing intestine at hatching and in skeletal muscle, skin and yolk sac. MSTN immunostaining was evident in several tissues and organs in both larvae and adults. Both IGF-I and MSTN proteins were detected in the liver from day 4 post-hatching and, subsequently, in the kidney and heart muscle from day 10. Our results suggest, on the basis of a combined methodological approach, that IGF-I and MSTN are involved in the regulation of somatic growth in the sea bass. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users. This research was supported by grants from the Italian Ministero dell’Università e della Ricerca Scientifica e Tecnologica (MIUR) and by the University of Padua (Progetto di Ateneo).     

哲罗鱼稚鱼最佳投喂策略

为探讨哲罗鱼稚鱼的最佳投喂策略,设置了饥饿再投喂试验、饥饿再投喂恢复试验以及日投喂频率试验.结果表明: 饥饿再投喂试验中,各饥饿组未表现出补偿生长现象.但在饥饿再投喂恢复试验中,各饥饿组表现出不同程度的补偿生长,其中S_1/2组(饥饿1/2 d投喂1/2 d)体质量的增加量与对照组接近,表现出完全补偿生长.表明在哲罗鱼早期稚鱼阶段(体质量0～2 g,水温9～15.3 ℃),S_1/2是可以考虑使用的投喂方法.日投喂频率试验中,T₃组(日投喂3次)体长、体质量的增加量以及特定生长率均最高,饵料转化率也相对较高.表明在哲罗鱼后期稚鱼阶段(体质量2～21 g,水温8.8～15.5 ℃),以日投喂3次为宜.     

Insulin-like growth factor I (IGF-I) in a growth-enhanced transgenic (GH-overexpressing) bony fish,the tilapia (<Emphasis Type="Italic">Oreochromis niloticus</Emphasis>): indication for a higher impact of autocrine/paracrine than of endocrine IGF-I

Several lines of growth hormone (GH)-overexpressing fish have been produced and analysed for growth and fertility parameters. However, only few data are available on the growth-promoting hormone insulin-like growth factor I (IGF-I) that mediates most effects of GH, and these are contradictory. Using quantitative real-time RT-PCR, radioimmunoassay, in situ hybridization, immunohistochemistry, and radiochromatography we investigated IGF-I and IGF binding proteins (IGFBPs) in an adult (17 months old) transgenic (GH-overexpressing) tilapia (Oreochromis niloticus). The transgenics showed an around 1.5-fold increase in length and an approximately 2.3-fold higher weight than the non-transgenics. Using radioimmunoassay, the serum IGF-I levels were lower (6.22 ± 0.75 ng/ml) in transgenic than in wild-type (15.01 ± 1.49 ng/ml) individuals (P = 0.0012). Radioimmunoassayable IGF-I in transgenic liver was 4.2-times higher than in wild-type (16.0 ± 2.21 vs. 3.83 ± 0.71 ng/g, P = 0.0017). No hepatocytes in wild-type but numerous hepatocytes in transgenic liver contained IGF-I-immunoreactivity. RT-PCR revealed a 1.4-times higher IGF-I mRNA expression in the liver of the transgenics (10.51 ± 0.82 vs. 7.3 ± 0.49 pg/μg total RNA, P = 0.0032). In correspondence, in situ hybridization showed more IGF-I mRNA containing hepatocytes in the transgenics. A twofold elevated IGF-I mRNA expression was determined in the skeletal muscle of transgenics (0.33 ± 0.02 vs. 0.16 ± 0.01 pg/μg total RNA, P < 0.0001). Both liver and serum of transgenics showed increased IGF-I binding. The increased IGFBP content in the liver may lead to retention of IGF-I, and/or the release of IGF-I into the circulation may be slower resulting in accumulation of IGF-I in the hepatocytes. Our results indicate that the enhanced growth of the transgenics likely is due to enhanced autocrine/paracrine action of IGF-I in extrahepatic sites, as shown here for skeletal muscle.     

哲罗鱼稚鱼最佳投喂策略

为探讨哲罗鱼稚鱼的最佳投喂策略,设置了饥饿再投喂试验、饥饿再投喂恢复试验以及日投喂频率试验.结果表明: 饥饿再投喂试验中,各饥饿组未表现出补偿生长现象.但在饥饿再投喂恢复试验中,各饥饿组表现出不同程度的补偿生长,其中S_1/2组(饥饿1/2 d投喂1/2 d)体质量的增加量与对照组接近,表现出完全补偿生长.表明在哲罗鱼早期稚鱼阶段(体质量0～2 g,水温9～15.3 ℃),S_1/2是可以考虑使用的投喂方法.日投喂频率试验中,T₃组(日投喂3次)体长、体质量的增加量以及特定生长率均最高,饵料转化率也相对较高.表明在哲罗鱼后期稚鱼阶段(体质量2～21 g,水温8.8～15.5 ℃),以日投喂3次为宜.     

Evolution of insulin-like growth factor I (IGF-I): structure and expression of an IGF-I precursor from Xenopus laevis

By means of a cloning strategy employing the polymerase chain reaction, we have isolated and characterized cDNAs for Xenopus laevis insulin-like growth factor I (IGF-I). These cDNAs encode a primary IGF-I translation product of 153 residues that demonstrates considerable amino acid sequence similarity with IGF-IA peptides from other species. Fifty-seven of 70 residues of the mature protein are identical among human, rat, chicken, and Xenopus IGF-I, while less amino acid conservation is found at the COOH-terminus (25/35 identities) or at the NH2-terminus (24/48 identities) of the precursor protein. Despite the lower degree of structural similarity at the NH2-terminus, in vitro studies of IGF-I biosynthesis and proteolytic processing support a conserved function for the atypically long 48 residue NH2-terminal signal sequence in directing the nascent IGF-I peptide through the secretory pathway. The 5'-untranslated region of Xenopus IGF-I mRNA matches the human, rat, and chicken sequences in greater than 90% of 279 nucleotides. IGF-I mRNAs from all four species encode a conserved upstream open reading frame of 14 amino acids starting 240-250 nucleotides 5' to the translation start site, suggesting a possible role for this region in modulating IGF-I gene expression. The X. laevis IGF-I gene is transcribed and processed into three mRNAs of 1.6, 2.1, and 3.0 kilobases in liver, and IGF-I mRNAs can be detected in liver, lung, heart, kidney, and peritoneal fat of adult animals. These studies demonstrate that both the IGF-I protein precursor and potential regulatory regions of IGF-I mRNA have been conserved during vertebrate evolution, and indicate that like several other polypeptide growth factors, IGF-I may be of fundamental importance in regulating specific aspects of growth and development in all vertebrates.     

Plasma insulin-like growth factor-I concentrations and growth in juvenile halibut (Hippoglossus hippoglossus): effects of photoperiods and feeding regimes

The effects of photoperiod and feeding regimes on plasma IGF-I levels and their relationship with growth rate of juvenile halibut (initial mean weight 364 g) were investigated by rearing fish under five different photoperiod regimes and two feeding regimes for 14 months. The entire photoperiod experiment was divided into 3 phases where the fish in each phase were exposed to either natural photoperiod (N), stimulated photoperiod with long day and short night (S) or continuous light (L). Thus, the following five photoperiod combinations were tested: a) Control group (NNN) b) Group 2A (NLN) c) Group 2B (NNL) d) Long day-natural group (SNN) e) Production group (LNN). In addition, the Control group was split into two parts and fed according to two different feeding regimes: a) Continuous fed group: Fish fed every day. b) Starvation/re-fed group: Fish were starved for 5 weeks and then re-fed for 10 weeks, and the treatment repeated during the whole experimental period. The analyses of IGF-I were performed from individually tagged fish in all groups in September 2005 and March 2006. In order to test how rapidly starvation affects circulating IGF-I levels samples were taken from the Starvation/re-fed group after a 10 days starvation (September) and immediately after 10 weeks of feeding (March). A significant relationship between IGF-I levels and individual growth in the preceding period and photoperiod and starvation treatment was found on both occasions. In conclusion, the present study indicates that plasma IGF-I levels are correlated to growth in Atlantic halibut, and affected by photoperiod treatment or compensatory growth during re-feeding. Correlation between individual growth rate and IGF-I levels was low, but significant, highlighting the complexity of how environmental factors affect the endocrine and physiological regulation of growth in fish.     

Identification of two insulin-like growth factor IIs in the Japanese eel, Anguilla japonica: cloning, tissue distribution, and expression after growth hormone treatment and seawater acclimation

To better understand the role of IGFs in Japanese eel, Anguilla japonica, we cloned insulin-like growth factor-II (IGF-II) cDNAs and examined their mRNA expression in several tissues. Two eel IGF-II cDNAs, eIGF-II-1 and eIGF-II-2, were cloned from the liver. A signal peptide and a mature peptide of both preproIGF-IIs were composed of 47 amino acids (aa) and 69 aa, but they differed at 17 aa and 13 aa, respectively. The E domain of eIGF-II-1 was 49 aa longer than that of eIGF-II-2, and differed at 22 aa within 52 aa. The highest eIGF-II-1 and II-2 mRNA levels were observed in the liver, with detectable levels also found in all tissues examined. The eIGF-II-1 mRNA levels in the liver, heart, and muscle were higher in females than in males, whereas those in the stomach and intestine were lower in the females. The eIGF-II-2 mRNA levels in the liver and swim-bladder were also higher in females than in males whereas those in the stomach, spleen, and intestine were lower in the females. The eIGF-II-1 mRNA levels in the liver were higher in large compared to small glass eels, while the eIGF-II-2 mRNA levels did not correlate with body weight. Both eIGF-II mRNA levels in the liver increased after eel GH treatments in vivo and in vitro. No differences in both eIGF-II mRNA levels were observed in the gills, liver, stomach and whole kidney between seawater- and freshwater-reared eels.     

Ghrelin receptor agonist GHRP-2 prevents arthritis-induced increase in E3 ubiquitin-ligating enzymes MuRF1 and MAFbx gene expression in skeletal muscle

Chronic arthritis is a catabolic state associated with an inhibition of the IGF system and a decrease in body weight. Cachexia and muscular wasting is secondary to protein degradation by the ubiquitin-proteasome pathway. The aim of this work was to analyze the effect of adjuvant-induced arthritis on the muscle-specific ubiquitin ligases muscle ring finger 1 (MuRF1) and muscle atrophy F-box (MAFbx) as well as on IGF-I and IGF-binding protein-5 (IGFBP-5) gene expression in the skeletal muscle. We also studied whether the synthetic ghrelin receptor agonist, growth hormone releasing peptide-2 (GHRP-2), was able to prevent arthritis-induced changes in the skeletal muscle. Arthritis induced an increase in MuRF1, MAFbx (P < 0.01), and tumor necrosis factor (TNF)-alpha mRNA (P < 0.05) in the skeletal muscle. Arthritis decreased the serum IGF-I and its gene expression in the liver (P < 0.01), whereas it increased IGF-I and IGFBP-5 gene expression in the skeletal muscle (P < 0.01). Administration of GHRP-2 for 8 days prevented the arthritis-induced increase in muscular MuRF1, MAFbx, and TNF-alpha gene expression. GHRP-2 treatment increased the serum concentrations of IGF-I and the IGF-I mRNA in the liver and in the cardiac muscle and decreased muscular IGFBP-5 mRNA both in control and in arthritic rats (P < 0.05). GHRP-2 treatment increased muscular IGF-I mRNA in control rats (P < 0.01), but it did not modify the muscular IGF-I gene expression in arthritic rats. These data indicate that arthritis induces an increase in the activity of the ubiquitin-proteasome proteolytic pathway that is prevented by GHRP-2 administration. The parallel changes in muscular IGFBP-5 and TNF-alpha gene expression with the ubiquitin ligases suggest that they can participate in skeletal muscle alterations during chronic arthritis.     

Distinct organ-specific up- and down-regulation of IGF-I and IGF-II mRNA in various organs of a GH-overexpressing transgenic Nile tilapia

Several lines of GH-overexpressing fish have been produced and characterized concerning organ integrity, growth, fertility and health but few and contradictory data are available on IGF-I that mediates most effects of GH. Furthermore, nothing is known on IGF-II. Thus, the expression of both IGFs in liver and various extrahepatic sites of adult transgenic (GH-overexpressing) tilapia and age-matched wild-type fish was determined by real-time PCR. Both IGF-I and IGF-II mRNA were found in all organs investigated and were increased in gills, kidney, intestine, heart, testes, skeletal muscle and brain of the transgenics (IGF-I: 1.4–4-fold; IGF-II: 1.7–4.2-fold). Except for liver, brain and testis the increase in IGF-I mRNA was higher than that in IGF-II mRNA. In pituitary, no significant change in IGF-I or IGF-II mRNA was detected. In spleen, however, IGF-I and IGF-II mRNA were both decreased in the transgenics, IGF-I mRNA even by the 19-fold. In agreement, in situ hybridisation revealed a largely reduced number of IGF-I mRNA-containing leukocytes and macrophages when compared to wild-type. These observations may contribute to better understanding the reported impaired health of GH-transgenic fish. Growth enhancement of the transgenics may be due to the increased expression of both IGF-I and IGF-II in extrahepatic sites. It is also reasonable that the markedly enhanced expression of liver IGF-II mRNA that may mimick an early developmental stage is a further reason for increased growth.     

Comparison of compensatory growth responses of juvenile three-spined stickleback and minnow following similar food deprivation protocols

The compensatory growth responses of individual juveniles of two co- existing species were compared after identical periods of starvation to determine inter-specific similarities and differences. The carnivorous stickleback Gasterosteus aculeatus was compared with the omnivorous minnow Phoxinus phoxinus. Both species experienced 1 or 2 weeks of starvation before being re-fed ad libitum. The two species differed in their response to the starvation periods, with minnows showing a lower weight-specific loss. Both species showed compensatory responses in appetite, growth and to a lesser extent, growth efficiency. Minnows wholly compensated for 1 and 2 weeks of starvation. At the end of the experiment, sticklebacks starved for 2 weeks were still showing a compensatory response and had not achieved full compensation. The compensatory responses of the sticklebacks showed a lag of a week before developing in the re-feeding phase, whereas the response of the minnows was immediate. Analysis of lipid and dry matter concentrations suggested that the compensatory response restored reserve lipids while also bringing the fish back to the growth trajectory of continuously fed fish.