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 growt

Insulin-like growth factor I (IGF-I) in a growth-enhanced transgenic (GH-overexpressing) bony fish, the tilapia ( Oreochromis niloticus ): 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±0.75ng/ml) in transgenic than in wild-type (15.01±1.49ng/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±0.71ng/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±0.49pg/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±0.01pg/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 muscl

Effects of seawater and freshwater challenges on the Gh/Igf system in the saline-tolerant blackchin tilapia (Sarotherodon melanotheron)

Prolactin (Prl) and growth hormone (Gh) as well as insulin-like growth factor 1 (Igf1) are involved in the physiological adaptation of fish to varying salinities. The Igfs have been also ascribed other physiological roles during development, growth, reproduction and immune regulation. However, the main emphasis in the investigation of osmoregulatory responses has been the endocrine, liver-derived Igf1 route and local regulation within the liver and osmoregulatory organs. Few studies have focused on the impact of salinity alterations on the Gh/Igf-system within the neuroendocrine and immune systems and particularly in a salinity-tolerant species, such as the blackchin tilapia Sarotherodon melanotheron. This species is tolerant to hypersalinity and saline variations, but it is confronted by severe climate changes in the Saloum inverse estuary. Here we investigated bidirectional effects of increased salinity followed by its decrease on the gene regulation of prl, gh, igf1, igf2, Gh receptor and the tumor-necrosis factor a. A mixed population of sexually mature 14-month old blackchin tilapia adapted to freshwater were first exposed to seawater for one week and then to fresh water for another week. Brain, pituitary, head kidney and spleen were excised at 4 h, 1, 2, 3 and 7 days after both exposures and revealed differential expression patterns. This investigation should give us a better understanding of the role of the Gh/Igf system within the neuroendocrine and immune organs and the impact of bidirectional saline challenges on fish osmoregulation in non-osmoregulatory organs, notably the complex orchestration of growth factors and cytokines

Somatostatin- und Gentamicin-induzierter Haarzellverlust

Einführung: Aminoglykoside können Haarzellen irreversibel schädigen. Somatostatin, ein Peptid mit Eigenschaften von einem Hormon/Neurotransmitter, zeigt einen neuroprotektiven Effekt durch die Bindung an seinen Rezeptor. Wir untersuchten, ob Somatostatin Haarzellen vor Gentamicin-induzierter Schädigung in vitro schützen kann. Material und Methoden: Mittels Reverse Transcriptase- Polymerase Chain Reaction (RT-PCR) wurde die Expression der Somatostatinrezeptor-mRNA in der Rattencochlea analysiert. Der Einfluss von Somatostatin auf die Gentamicin-induzierte Haarzellschädigung in vitro wurde mittels zweier verschiedener Konzentrationen (1 mM und 5 mM) Somatostatin geprüft. Resultate: Wir konnten mittels RT-PCR Somatostatinrezeptor- 1- und Somatostatinrezeptor-2-mRNA im Cortischen Organ, im Spiralganglion und in der Stria vascularis nachweisen. Des Weiteren konnten wir eine signifikante Abnahme der Anzahl abgestorbener Haarzellen bei mit entweder 1 mM oder 1 mM Somatostatin vorbehandelten Cortischen Organen im Vergleich zu Cortischen Organen, welche mit Gentamicin allein behandelt wurden, feststellen. Diskussion: Die Abnahme des Gentamicin-induzierten Haarzellverlustes durch die Vorbehandlung der Cortischen Organe durch Somatostatin ist ein Hinweis für einen potektiven Effekt von Somatostatin auf den Gentamicin-induzierten Haarzelverlust in vitro

Stress and survival pathways in the mammalian cochlea

Studies conducted over the last few years demonstrated that signaling pathways that operate in the organs of Corti (OC) play a central role in survival and death of hair cells. An important goal of molecular otology is to characterize these signaling pathways in normal inner ears and inner ears exposed to a variety of different forms of stress, such as ototoxic substances and noise overexposure. In this study, we used high-performance reverse protein microarray technology and phospho-specific antibodies to examine the activation status of defined molecules involved in cellular signaling. We demonstrate that reverse protein microarrays based on the highly sensitive planar-waveguide technology provide an effective and high-throughput means to assess the activation state of key molecules involved in apoptotic and prosurvival signaling in microdissected OC explants over time. In this study, we show that gentamicin and a specific NF-kappaB inhibitor increase the ratio of phospho-c-Jun/c-Jun in OC explants of postnatal rats soon after exposure to these drugs. In addition, we found a decrease in the phospho-Akt/Akt ratio in OC explants early after NF-kappaB inhibition. Finally, we observed an early and consistent decrease in the phospho-p38/p38 ratio in OC explants exposed to the NF-kappaB inhibitor and only a transient decrease in this ratio in OC examples after gentamicin exposure