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. GenotypeEnvironmental factors Growth Food

3. Growth hormone (GH) transgenic animals

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

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

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

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

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

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

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

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

12. The hepatic expression IGF-1 is under the dual regulation of nutrients and GH of nutrients and GH IGFs GH IGFs Extra-hepatic tissues Liver Brain Environment 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. pituitary

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

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

15. Low O 2 Normal O 2 IGFBP-1 is a hypoxia-inducible gene in zebrafish fish embryos and larvae Kajimura et al. PNAS, 2005 96 hpf

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

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 2 month-old fish were fed with various rations (1, 3, and 9% of body weight) for 4 weeks 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 4 fish were sacrificed and their IGFBP-1a/-1b mRNA levels were measured by qRT-PCR and normalized by β-actin mRNA levels At each sampling time, 20 fish were randomly picked up to measure their body length and body weight at each time point 40-50 fish in each experimental group Each experiment has two parallel groups

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

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

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

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

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

24. Conclusions and Discussion 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. The GH-IGF-I axis plays central roles in fish growth regulation. 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. IGFs in blood and other extracellular fluids are present in complexes with several IGFBPs. The levels of IGFBP-1 expression is reversely correlated with growth. IGFBP-1 is highly induced by food deprivation, malnutrition, stress, and hypoxia.

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

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