Cunming Duan University of Michigan, Ann Arbor, MI

1. Regulation of IGF and IGF binding protein gene expression by nutrition and aquaculture related stressors: Implications in fish growth and stress assessment Cunming Duan University of Michigan, Ann Arbor, MI

2. Genotype Environmental factors Food Growth

3. Growth hormone (GH) transgenic animals

4. "Paracrine, autocrine" IGFs "Endocrine" The growth-promoting action of GH in postnatal stages is mediated by Insulin-like growth factors (IGFs) Environment "Somatomedin Hypothesis" Brain pituitary - GH + Liver + Extra-hepatic tissues (somatic, gonadal) IGFs Duan (1997) Am. Zool.

5. Insulin-like Growth Factor-1 S S S G R R Y A G P C-domain T Q P T D-domain G K A S K I N A V D P F L G C R K D Y S R P E L L F R E E C P A-domain M G C F T Y A R L C C D G G A B-domain C E V L V F D A L Q

6. Insulin-like Growth Factor (IGF) Signaling: IGF-1 IGF-2 IGF1R Growth Development Reproduction Aging

7. Woods et al., N Engl J Med, 1996 Abuzzahab et al., N Engl J Med, 2003 Sutter et al., Science, 2007 Liu et al., Cell, 1993 Baker et al., Cell, 1993 Control IGF-IR deficient Walenkamp et al., JCEM, 2005 Schlueter et al., FASEB J, 2006 The IGF signaling is a evolutionarily conserved pathway that plays fundamental roles in growth control

8. "Paracrine, autocrine" IGFs "Endocrine" IGFs mediate GH actions in postnatal stages Environment Brain pituitary - GH + Liver + Extra-hepatic tissues (somatic, gonadal) IGFs

9. J. Nutrition, 1998 _ • + elevated levels • reduced levels • ? unknown

10. Starvation Undernourishment Malnourishment IGF-1 expression is also regulated by nutritional factors Environment Brain pituitary GH IGFs Liver Extra-hepatic tissue growth (muscle, bone, gonad etc.) IGFs Wood, A.W., Duan, C., and Bern, H.A. (2005). Insulin-like growth factor signaling in fish. Int. Rev. Cyto. 243: 215-285

11. IGF-1 expression is also regulated by nutritional factors Environment Brain pituitary GH IGFs Starvation Undernourishment Malnourishment Liver Extra-hepatic tissue growth (muscle, bone, gonad etc.) IGFs Wood, A.W., Duan, C., and Bern, H.A. (2005). Insulin-like growth factor signaling in fish. Int. Rev. Cyto. 243: 215-285

12. Environment Brain GH IGFs Liver Nutrients • The nutritional regulation of the GH-IGF-I axis is an interface between nutrients and hormones acting in concert to control animal growth. • This interface is conserved throughout vertebrate evolution. Extra-hepatic tissues IGFs The hepatic expression IGF-1 is under the dual regulation of nutrients and GH pituitary

13. ALS BP BP IGF IGF IGF • Prolong the half-life of IGFs. • Regulate the biological activities of IGFs by controlling their access to the IGF-1 receptors on target cells. IGFs are bound to IGF binding proteins (IGFBP) in extracellular fluids

14. Hypoxia Normoxia IGFBP-1 100 * IGFBP-1 mRNA/ß-actin mRNA (fold) 10 1 .1 Hypoxia Normoxia Maures and Duan, 2002, Endocrinology IGFBP-1 is induced by stressors Kelley et al., 2002, J. Endocrinology

15. IGFBP-1 is a hypoxia-inducible gene in zebrafish fish embryos and larvae Low O2 Normal O2 96 hpf Kajimura et al. PNAS, 2005

16. (3) Reduced IGF signaling (4) Suppression of IGF actions IGFBP-1 is a metabolically controlled molecular switch that inhibits IGF actions under catabolic states Catabolic or stressful conditions (1) Induction of IGFBP-1 BP-1 BP-1 BP-1 BP-1 (5) Growth retardation Developmental delay (2) Reduced “available” IGF IGF receptor Kajimura et al., PNAS (2005); Kajimura and Duan Fish Biol. (2007)

17. Fasting induces and re-feeding reduces IGFBP-1 gene expression in young zebrafish Kamei, Lu, Jiao, et al., PLoS One, 2008

18. Experimental Design 40-50 fish in each experimental group 2 month-old fish were fed with various rations (1, 3, and 9% of body weight) for 4 weeks At each sampling time, 20fish were randomly picked up to measure their body length and body weight at each time point 4 fish were sacrificed and their IGFBP-1a/-1b mRNA levels were measured by qRT-PCR and normalized by β-actin mRNA levels 1-month old juvenile fish were fed with different diets.They are standard diet followed with: high vs. low carbohydrate; high vs. low protein; high vs. low fat; high vs. low vitamin contents Each experiment has two parallel groups

19. Effects of different feeding rations on growth and IGFBP-1a and -1b gene expressions a a b bc c c d de fg fgh fgh ef fg gh gh gh gh Feng Q et al. unpublished data

20. IGFBP-1a a ab abc abcd abcd abcd abcd abcd abcd abcd abcd abcd cd abcd bcd d IGFBP-1b a ab abc bcd abcd bcd cde bcde bcd bcde bcd bcde de bcde bcde e Feng Q et al. unpublished data

21. Effect of diet protein levels on growth and IGFBP-1 gene expression a a b cd bc d d e e ef g efg efg Feng et al. unpublished data

22. IGFBP-1a a a a ab bc bcd bc cde cde bcd de cde e IGFBP-1b a ab ab ab ab ab abc abc abc de cde e de Feng et al. unpublished data

23. IGFBP-1a a ab abc abcd abcd bcdef bcde cdef def ef bcdef f ef IGFBP-1b a ab abc ab abc abc abc bcd cd cd cd cd d Feng et al. unpublished data

24. Conclusions and Discussion • The GH-IGF-I axis plays central roles in fish growth regulation. • The hepatic IGF-I expression is under the regulation of GH and nutritional states. This dual regulation of IGF-I represents an interface between nutrients and hormones acting in concert to control animal growth. • IGFs in blood and other extracellular fluids are present in complexes with several IGFBPs. • IGFBP-1 is highly induced by food deprivation, malnutrition, stress, and hypoxia. • The levels of IGFBP-1 expression is reversely correlated with growth. • These results suggest that the levels of IGF-1 and IGFBP-1 expression (or their ratio) are useful indicators of growth and may be useful tools in nutrient utilization and stress assessment.

25. Hypoxia followed by re-oxygenation accelerates growth and developmental rates Catch-up growth Kamei et al. (2011) Development

27. National Heart, Lung and Blood Institute , NIH Supported by grants from: National Science FoundationNational Institutes of Health