Supplementary Material for: Cotreatment of IGF1 and Fadrozole Upregulates the Expression of RSPO1, SOX9, and AMH in Chicken Embryos

Insulin-like growth factor-1 (IGF1) and anti-aromatase synergistically increase the rate and stability of female-to-male sex reversal as well as pre- and postnatal weight gains in hatched chickens. This study aimed at assessing gene expression profiles of chicken embryos treated with IGF1 and fadrozole. Day 3.5 fertile eggs were in ovo injected with one of IGF1, fadrozole anti-aromatase, combined IGF1 and fadrozole, or sham injection. The expression profile was studied on day 6 and day 11 of the embryonic development following gonadal differentiation. On day 6 of embryonic development, simultaneous injection of IGF1 and fadrozole significantly upregulated the expression of RSPO1, AMH, and SOX9 in genetically female embryos compared to single injections and control groups. Also, a higher expression of ESR1 and BMP4 was observed in genetically male embryos on day 6 compared to the control group. In day 11 embryos, a higher expression of BMP4 was detected in both males and females of the IGF1 and fadrozole-administered group compared to the sham injection cohort. In conclusion, the results of this study indicate that combined effects of IGF1 and fadrozole induce female-to-male sex reversal by increasing the expression of testis developmental factors rather than attenuating ovary developmental factors.