Transgenic Research

, Volume 16, Issue 4, pp 479–489 | Cite as

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

  • Elisabeth Eppler
  • Antje Caelers
  • Natallia Shved
  • Guylin Hwang
  • Azizur M. Rahman
  • Norman Maclean
  • Jürgen Zapf
  • Manfred Reinecke
Original Paper


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.


IGF-I IGFBP Liver Serum Skeletal muscle Transgenic fish 



The authors are grateful to Ms Cornelia Zwimpfer, Division of Endocrinology and Diabetes, Department of Medicine, University Hospital Zürich, for excellent support in the performance of radioimmunoassay and radiochromatography. The study was supported by the Swiss National Science Foundation (grant no. 111028) and the Hartmann Müller-Foundation for Medical Research (grant no. 1115).


  1. Amsterdam A, Becker TS (2005) Transgenes as screening tools to probe and manipulate the zebrafish genome. Dev Dyn 234:255–268PubMedCrossRefGoogle Scholar
  2. Baños N, Planas JV, Gutiérrez J, Navarro I (1999) Regulation of plasma insulin-like growth factor-I levels in brown trout (Salmo trutta). Comp Biochem Physiol C Pharmacol Toxicol Endocrinol 124:33–40PubMedCrossRefGoogle Scholar
  3. Baxter RC, Martin JL, Beniac V (1989) High molecular weight insulin-like growth factor binding protein complex. Purification and properties of the acid-labile subunit from human serum. J Biol Chem 264:11843–11848PubMedGoogle Scholar
  4. Beckman BR, Larsen DA, Moriyama S, Lee-Pawlak B, Dickhoff WW (1998) Insulin-like growth factor-I and environmental modulation of growth during smoltification of spring chinook salmon (Oncorhynchus tshawystscha). Gen Comp Endocrinol 109:325–335PubMedCrossRefGoogle Scholar
  5. Berishvili G, D’Cotta H, Baroiller J-F, Segner H, Reinecke M (2006a) Differential expression of IGF-I mRNA and peptide in the male and female gonad during early development of a bony fish, the tilapia Oreochromis niloticus. Gen Comp Endocrinol 146:204–210CrossRefGoogle Scholar
  6. Berishvili G, Shved N, Eppler E, Clotà F, Baroiller JF, Reinecke M (2006b) Organ-specific expression of IGF-I during early development of bony fish as revealed in the tilapia, Oreochromis niloticus, by in situ hybridization and immunohistochemistry: indication for the particular importance of local IGF-I. Cell Tissue Res 325:287–301CrossRefGoogle Scholar
  7. Butler A, LeRoith D (2001) Minireview: tissue-specific versus generalized gene targeting of the igf1 and igf1r genes and their roles in insulin-like growth factor physiology. Endocrinology 142:1685–1688PubMedCrossRefGoogle Scholar
  8. Caelers A, Berishvili G, Meli ML, Eppler E, Reinecke M (2004) Establishment of a real-time RT-PCR for the determination of absolute amounts of IGF-I and IGF-II gene expression in liver and extrahepatic sites of the tilapia. Gen Comp Endocrinol 137:196–204PubMedCrossRefGoogle Scholar
  9. Caelers A, Mclean N, Hwang G, Eppler E, Reinecke M (2005) Expression of endogenous and exogenous growth hormone (GH) messenger (m) RNA in a GH-transgenic tilapia (Oreochromis niloticus). Transgenic Res 14:95–104PubMedCrossRefGoogle Scholar
  10. Devlin RH, Vesaki TY, Biagi CA, Donaldson EM, Swanson P, Chan W-K (1994) Extraordinary salmon growth. Nature 371:209–210CrossRefGoogle Scholar
  11. Devlin RH, Swanson P, Clarke WC, Plisetskaya E, Dickhoff W, Moriyama S, Yesaki TY, Hew C-L (2000) Seawater adaptability and hormone levels in growth-enhanced transgenic coho salmon, Oncorhynchus kisutch. Aquaculture 191:367–385CrossRefGoogle Scholar
  12. Du SJ, Gong ZY, Fletcher GL, Shears MA, King MJ, Idler DR, Hew CL (1992) Growth enhancement in transgenic Atlantic salmon by the use of an “all fish” chimeric growth hormone gene construct. Biotechnology (NY) 10:176–181CrossRefGoogle Scholar
  13. Duan C (1998) Nutritional and developmental regulation of insulin-like growth factors in fish. J Nutr 128(Suppl 2):306S–314SPubMedGoogle Scholar
  14. Duan C, Xu Q (2005) Roles of insulin-like growth factor (IGF) binding proteins in regulating IGF actions. Gen Comp Endocrinol 142:44–52PubMedCrossRefGoogle Scholar
  15. Duan C, Duguay SJ, Plisetskaya EM (1993a) Insulin-like growth factor (IGF-I) mRNA expression in coho salmon, Oncorhynchus kisutch: tissue distribution and effects of growth hormone/prolactin family proteins. Fish Physiol Biochem 11:371–379CrossRefGoogle Scholar
  16. Duan C, Hanzawa N, Takeuchi Y, Hamada E, Miyachi S, Hirano T (1993b) Use of primary cultures of salmon hepatocytes for the study of hormonal regulation of IGF-I expression in vitro. Zool Sci 10:473–480Google Scholar
  17. Duguay SJ, Lai-Zhang J, Steiner DF, Funkenstein B, Chan SJ (1996) Developmental and tissue-regulated expression of IGF-I and IGF-II mRNAs in Sparus aurata. J Mol Endocrinol 16:123–132PubMedGoogle Scholar
  18. Dyer AR, Upton Z, Stone D, Thomas PM, Soole KL, Higgs N, Quinn K, Carragher JF (2004) Development and validation of a radioimmunoassay for fish insulin-like growth factor I (IGF-I) and the effect of aquaculture related stressors on circulating IGF-I levels. Gen Comp Endocrinol 135:268–275PubMedCrossRefGoogle Scholar
  19. Fiess JC, Kunkel-Patterson A, Mathias L, Riley LG, Yancey PH, Hirano T, Grau EG (2007) Effects of environmental salinity and temperature on osmoregulatory ability, organic osmolytes, and plasma hormone profiles in the Mozambique tilapia (Oreochromis mossambicus). Comp Biochem Physiol A Mol Integr Physiol 146:252–264PubMedCrossRefGoogle Scholar
  20. Gabillard JC, Weil C, Rescan PY, Navarro I, Gutierrez J, Le Bail PY (2003) Effects of environmental temperature on IGF1, IGF2, and IGF type I receptor expression in rainbow trout (Oncorhynchus mykiss). Gen Comp Endocrinol 133:233–242PubMedCrossRefGoogle Scholar
  21. Gray ES, Kelley KM, Law S, Tsai R, Young G, Bern H (1992) Regulation of hepatic growth hormone receptors in coho salmon (Oncorhynchus kisutch). Gen Comp Endocrinol 88:243–252PubMedCrossRefGoogle Scholar
  22. Guillén II, Lleonart R, Agramonte A, Morales R, Morales A, Hernandez CA, Vazquez MM, Diaz M, Herrera MT, Alvarez-Lajonchere L, Hernandez O, de la Fuente J (1998) Physiological changes in the juvenile euryhaline teleost, the tilapia Oreochromis hornorum, injected with E. coli-derived homologous growth hormone. J Mar Biotechnol 6:142–151PubMedGoogle Scholar
  23. Hansson H-A, Nilsson A, Isgaard J, Billig H, Isaksson O, Skottner A, Andersson IK, Rozell B (1988) Immunohistochemical localization of insulin-like growth factor I in the adult rat. Histochemistry 89:403–410PubMedCrossRefGoogle Scholar
  24. Hwang GL, Rahman MA, Abdul Razak S, Sohm F, Farahmand H, Smith A, Brooks C, Maclean N (2003) Isolation and characterisation of tilapia beta-actin promoter and comparison of its activity with carp beta-actin promoter. Biochim Biophys Acta 1625:11–18PubMedGoogle Scholar
  25. Jones JI, Clemmons DR (1995) Insulin-like growth factors and their binding proteins: biological actions. Endocr Rev 16:3–34PubMedCrossRefGoogle Scholar
  26. Kajimura S, Uchida K, Yada T, Riley LG, Byatt JC, Colloier RJ, Aida K, Hirano T, Grau EG (2001) Stimulation of insulin-like growth factor-I production by recombinant bovine growth hormone in Mozambique tilapia, Oreochromis mossambicus. Fish Physiol Biochem 25:221–230CrossRefGoogle Scholar
  27. Kelley KM, Siharath K, Bern HA (1992) Identification of insulin-like growth factor-binding proteins in the circulation of four teleost fish species. J Exp Zool 263:220–224PubMedCrossRefGoogle Scholar
  28. Kelley KM, Schmidt KE, Berg L, Sak K, Galima MM, Gillespie C, Balogh L, Hawayek A, Reyes JA, Jamison M (2002) Comparative endocrinology of the insulin-like growth factor-binding protein. J Endocrinol 175:3–18PubMedCrossRefGoogle Scholar
  29. Kelley KM, Price TD, Galima MM, Sak K, Reyes JA, Zepeda O, Hagstrom R, Tuan A, Truong TA, Lowe CG (2006) Insulin-like growth factor binding proteins (IGFBPs) in fish: beacons for (disrupted) growth endocrine physiology. In: Reinecke M, Zaccone G, Kapoor BG (eds) Fish endocrinolgy. Science Publishers, Enfield, pp 167–195Google Scholar
  30. Larsen DA, Beckman BR, Dickhoff WW (2001) The effect of low temperature and fasting during the winter on metabolic stores and endocrine physiology (insulin, insulin-like growth factor-I, and thyroxine) of coho salmon, Oncorhynchus kisutch. Gen Comp Endocrinol 123:308–323PubMedCrossRefGoogle Scholar
  31. Lu JK, Fu BH, Wu JL, Chen TT (2002) Production of transgenic silver sea bream (Sparus sarba) by different gene transfer methods. Mar Biotechnol (NY) 4:328–337CrossRefGoogle Scholar
  32. Maclean N, Rahman MA, Sohm F, Hwang G, Iyengar A, Ayad H, Smith A, Farahmand H (2002) Transgenic tilapia and the tilapia genome. Gene 295:265–277PubMedCrossRefGoogle Scholar
  33. Martinez R, Estrada MP, Berlanga J, Guillén I, Hernandez O, Cabrera E, Pimentel R, Morales R, Herrera F, Morales A, Pina JC, Abad Z, Sanchez V, Melamed P, Lleonart R, de la Fuente J (1996) Growth enhancement in transgenic tilapia by ectopic expression of tilapia growth hormone. Mol Mar Biol Biotechnol 5:62–70PubMedGoogle Scholar
  34. McCormick SD, Kelley KM, Young G, Nishioka RS, Bern HA (1992) Stimulation of coho salmon growth by insulin-like growth factor I. Gen Comp Endocrinol 86:398–406PubMedCrossRefGoogle Scholar
  35. Mingarro M, Vega-Rubin de Celis S, Astola A, Pendon C, Valdivia MM, Perez-Sanchez J (2002) Endocrine mediators of seasonal growth in gilthead sea bream (Sparus aurata): the growth hormone and somatolactin paradigm. Gen Comp Endocrinol 128:102–111PubMedCrossRefGoogle Scholar
  36. Moriyama S (1995) Increased plasma insulin-like growth factor-I (IGF-I) following oral and intraperitoneal administration of growth hormone to rainbow trout, Oncorhynchus mykiss. Growth Regul 5:164–167PubMedGoogle Scholar
  37. Moriyama S, Ayson FG, Kawauchi H (2000) Growth regulation by insulin-like growth factor-I in fish. Biosci Biotechnol Biochem 64:1553–1562PubMedCrossRefGoogle Scholar
  38. Ng TB, Leung TC, Cheng CHK, Woo NY (1992) Growth hormone binding sites in tilapia (Oreochromis mossambicus) liver. Gen Comp Endocrinol 86:111–118PubMedCrossRefGoogle Scholar
  39. Park R, Shepherd BS, Nishioka RS, Grau EG, Bern HA (2000) Effects of homologous pituitary hormone treatment on serum insulin-like growth factor-binding proteins (IGFBPs) in hypophysectomized tilapia, Oreochromis mossambicus, with special reference to a novel 20-kDa IGFBP. Gen Comp Endocrinol 117:404–412PubMedCrossRefGoogle Scholar
  40. Pierce AL, Beckman BR, Shearer KD, Larsen DA, Dickhoff WW (2001) Effects of ration on somatotropic hormones and growth in coho salmon. Comp Biochem Physiol B Biochem Mol Biol 128:255–264PubMedCrossRefGoogle Scholar
  41. Pierce AL, Fukada H, Dickhoff WW (2005) Metabolic hormones modulate the effect of growth hormone (GH) on insulin-like growth factor-I (IGF-I) mRNA level in primary culture of salmon hepatocytes. J Endocrinol 184:341–349PubMedCrossRefGoogle Scholar
  42. Plisetskaya EM (1998) Some of my not so favorite things about insulin and insulin-like growth factors in fish. Comp Biochem Physiol B Biochem Mol Biol 121:3–11PubMedCrossRefGoogle Scholar
  43. Rahman A, Maclean N (1992) Fish transgene expression by direct injection into fish muscle. Mol Mar Biol Biotechnol 1:286–289PubMedGoogle Scholar
  44. Rahman MA, Mak R, Ayad H, Smith A, Maclean N (1998) Expression of a novel piscine growth hormone gene results in growth enhancement in transgenic tilapia (Oreochromis niloticus). Transgenic Res 7:357–369PubMedCrossRefGoogle Scholar
  45. Reinecke M, Collet C (1998) The phylogeny of the insulin-like growth factors. Int Rev Cytol 183:1–94PubMedCrossRefGoogle Scholar
  46. Reinecke M, Schmid A, Ermatinger R, Loffing-Cueni D (1997) Insulin-like growth factor I in the teleost Oreochromis mossambicus, the tilapia: gene sequence, tissue expression, and cellular localization. Endocrinology 138:3613–3619PubMedCrossRefGoogle Scholar
  47. Reinecke M, Björnsson TB, Dickhoff WW, McCormick SD, Navarro I, Power DM, Gutiérrez J (2005) Growth hormone and insulin-like growth factors in fish: where we are and where to go. Gen Comp Endocrinol 142:20–24PubMedCrossRefGoogle Scholar
  48. Richardson NA, Anderson AJ, Rimmer MA, Sara VR (1995) Localization of insulin-like growth factor-I immunoreactivity in larval and juvenile barramundi (Lates calcarifer). Gen Comp Endocrinol 100:282–292PubMedCrossRefGoogle Scholar
  49. Riley LG, Richman NH 3rd, Hirano T, Grau EG (2002) Activation of the growth hormone/insulin-like growth factor axis by treatment with 17 alpha-methyltestosterone and seawater rearing in the tilapia, Oreochromis mossambicus. Gen Comp Endocrinol 127:285–292PubMedCrossRefGoogle Scholar
  50. Rocha A, Ruiz S, Estepa A, Coll JM (2004) Application of inducible and targeted gene strategies to produce transgenic fish: a review. Mar Biotechnol (NY) 6:118–127Google Scholar
  51. Schmid AC, Naef E, Kloas W, Reinecke M (1999) IGF-I and IGF-II in the ovary of a bony fish Oreochromis mossambicus, the tilapia: In situ hybridization, immunohistochemical localisation, Northern Blot and cDNA sequences. Mol Cell Endocrinol 156:141–149PubMedCrossRefGoogle Scholar
  52. Schmid AC, Reinecke M, Kloas W (2000) Primary cultured hepatocytes of the bony fish, Oreochromis mossambicus, the tilapia: a valid tool for physiological studies on IGF-I expression in liver. J Endocrinol 166:265–273PubMedCrossRefGoogle Scholar
  53. Shamblott MJ, Cheng CM, Bolt D, Chen TT (1995) Appearance of insulin-like growth factor mRNA in the liver and pyloric ceca of a teleost in response to exogenous growth hormone. Proc Natl Acad Sci USA 92:6943–6946PubMedCrossRefGoogle Scholar
  54. Shepherd BS, Sakamoto T, Nishioka RS, Richman NH III, Mori I, Madsen SS, Chen TT, Hirano T, Bern HA, Grau EG (1997) Somatotrophic actions of homologous growth hormone and prolactins in the euryhyaline teleost, the tilapia, Oreochromis mossambicus. Proc Natl Acad Sci USA 94:2068–2072PubMedCrossRefGoogle Scholar
  55. Shimizu M, Swanson P, Dickhoff WW (1999) Free and protein-bound insulin-like growth factor-I (IGF-I) and IGF-binding proteins in plasma of coho salmon, Oncorhynchus kisutch. Gen Comp Endocrinol 115:398–405PubMedCrossRefGoogle Scholar
  56. Shimizu M, Swanson P, Fukuda H, Hara A, Dickhoff WW (2000) Comparison of extraction methods and assay validation for salmon insulin-like growth factor-I using commercially available components. Gen Comp Endocrinol 119:26–36PubMedCrossRefGoogle Scholar
  57. Shimizu M, Hara A, Dickhoff WW (2003) Development of an RIA for salmon 41 kDa IGF-binding protein. J Endocrinol 178:275–283PubMedCrossRefGoogle Scholar
  58. Shimizu M, Dickey JT, Fukada H, Dickhoff WW (2005) Salmon serum 22 kDa insulin-like growth factor-binding protein (IGFBP) is IGFBP-1. J Endocrinol 184:267–276PubMedCrossRefGoogle Scholar
  59. Siharath K, Nishioka RS, Madsen SS, Bern HA (1995) Regulation of IGF-binding proteins by growth hormone in the striped bass, Morone saxatilis. Mol Mar Biol Biotechnol 4:171–178Google Scholar
  60. Sjögren K, Liu J-L, Blad K, Skrtic S, Vidal O, Wallenius V, Le Roith D, Törnell J, Isaksson OPG, Jansson J-O, Ohlsson C (1999) Liver-derived insulin-like growth factor I (IGF-I) is the pricipal source of IGF-I in blood but is not required for postnatal body growth in mice. Proc Natl Acad Sci USA 96:7088–7092PubMedCrossRefGoogle Scholar
  61. Sundstrom LF, Lohmus M, Johnsson JI, Devlin RH (2004) Growth hormone transgenic salmon pay for growth potential with increased predation mortality. Proc Biol Sci 271(Suppl 5):S350–S352PubMedCrossRefGoogle Scholar
  62. Uchida K, Kajimura S, Riley LG, Hirano T, Aida K, Grau EG (2003) Effects of fasting on growth hormone/insulin-like growth factor I axis in the tilapia, Oreochromis mossambicus. Comp Biochem Physiol A Mol Integr Physiol 134:429–433PubMedCrossRefGoogle Scholar
  63. Vong QP, Chan KM, Cheng CH (2003) Quantification of common carp (Cyprinus carpio) IGF-I and IGF-II mRNA by real-time PCR: differential regulation of expression by GH. J Endocrinol 178:513–521PubMedCrossRefGoogle Scholar
  64. Wood AW, Duan C, Bern HA (2005) Insulin-like growth factor signaling in fish. Int Rev Cytol 243:215–285PubMedCrossRefGoogle Scholar
  65. Yakar S, Liu J-L, Stannard B, Butler A, Accili D, Sauer B, LeRoith D (1999) Normal growth and development in the absence of hepatic insulin-like growth factor I. Proc Natl Acad Sci USA 96:7324–7329PubMedCrossRefGoogle Scholar
  66. Zapf J (1995) Physiological role of the insulin-like growth factor binding proteins. Eur J Endocrinol 132:645–654PubMedCrossRefGoogle Scholar
  67. Zapf J, Waldvogel M, Froesch ER (1975) Binding of nonsuppressible insulin-like activity to human serum. Arch Biochem Biophys 168:638–645PubMedCrossRefGoogle Scholar
  68. Zapf J, Hauri C, Waldvogel M, Futo E, Häsler H, Binz K, Guler HP, Schmid C, Froesch ER (1989) Recombinant human insulin-like growth factor I induces its own specific carrier protein in hypophysectomized and diabetic rats. Proc Natl Acad Sci USA 86:3813–3817PubMedCrossRefGoogle Scholar
  69. Zbikowska HM (2003) Fish can be first—advances in fish transgenesis for commercial applications. Transgenic Res 12:379–389PubMedCrossRefGoogle Scholar
  70. Zhang PJ, Hayat M, Joyce C, Gonzalez-Villasenor LI, Lin CM, Dunham RA, Chen TT, Powers DA (1990) Gene transfer, expression and inheritance of pRSV-rainbow trout-GH cDNA in the common carp, Cyprinus carpio (Linnaeus). Mol Reprod Dev 25:3–13PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media B.V. 2007

Authors and Affiliations

  • Elisabeth Eppler
    • 1
  • Antje Caelers
    • 1
  • Natallia Shved
    • 1
  • Guylin Hwang
    • 2
  • Azizur M. Rahman
    • 2
  • Norman Maclean
    • 2
  • Jürgen Zapf
    • 1
  • Manfred Reinecke
    • 1
  1. 1.Division of Neuroendocrinology, Institute of AnatomyUniversity of ZürichZürichSwitzerland
  2. 2.School of Biological SciencesUniversity of SouthamptonSouthamptonUK

Personalised recommendations