Abstract
Graves' orbitopathy (GO), an extrathyroidal manifestation of Graves' disease, is an inflammatory autoimmune disorder of the orbit that involves the differentiation of precursor cells into mature adipocytes and retro-orbital adipose tissue accumulation. Here, we examined the involvement of autophagy in adipogenesis and explored the effects of icariin, a flavonoid isolated from the genus Epimedium with a wide range of biological and pharmacological effects, on autophagy and adipogenesis in 3T3-L1 preadipocytes and in a mouse model of GO. Microscopic examination of autophagosome formation and lipid droplet accumulation by Oil Red O staining, and western blot assessment of autophagic markers in the presence of the autophagy inhibitors Asn and 3-MA showed that autophagy is essential for adipogenesis. Icariin inhibited the differentiation of preadipocytes into mature adipocytes by suppressing autophagy, and these effects were mediated by the inhibition of AMPK/mTOR pathway activation. In a mouse model of thyroid stimulating hormone receptor induced GO, icariin reduced orbital muscle adipose tissue expansion and lipid droplet accumulation by inhibiting AMPK/mTOR mediated autophagy. Collectively, these results reveal a potential mechanism underlying the protective effects of icariin against autophagy induced adipogenesis and suggest that icariin could be developed as a new therapeutic candidate for the prevention and treatment of GO.
Highlights
Graves’ disease is an autoimmune disease characterized by the binding of autoantibodies to the thyrotropin receptor on follicular cells of the thyroid gland, resulting in the activation and enlargement of the thyroid gland (Davies et al, 2005)
The results showed that there was no significant difference between cell numbers with or without icariin indicating that it had no effect on cell viability (Figure S1A) or apoptosis rates (Figure S1B)
Consistent with the in vitro findings, analysis of orbital tissues by western blotting showed that the levels of phospho-mammalian target of rapamycin (mTOR) and phospho-AMPK were significantly increased in Graves’ orbitopathy (GO) mice, whereas icariin treatment reversed this effect (Figures 4E,G,H). These results indicated that icariin inhibits orbital muscle adipogenesis in a mouse model of GO, and this effect may be mediated by the suppression of autophagy through the AMPK signaling pathway
Summary
Graves’ disease is an autoimmune disease characterized by the binding of autoantibodies to the thyrotropin receptor on follicular cells of the thyroid gland, resulting in the activation and enlargement of the thyroid gland (Davies et al, 2005). Adipogenesis, the differentiation of pre-adipocytes into mature adipocytes, is characterized by the activation of transcription factors such as peroxisome proliferator-activated receptor γ, members of the CCAAT/enhancer-binding protein family, and sterol regulatory element binding protein-1 (Tang and Lane, 2012). These factors modulate the expression of genes essential for adipocyte differentiation, which is characterized by the formation of lipid droplets in the cytosol for energy storage (Zhang et al, 2012). Adipose tissue formation involves the process of autophagy, in which cytoplasmic materials are packaged into double membrane structures known as autophagosomes and targeted for lysosomal degradation (Singh et al, 2009). AMPK is activated by changes in the availability of ATP and AMP in the cytosol, and AMPK activation leads to the inactivation of energy consuming pathways and the activation of energy producing pathways, restoring energy homeostasis (Hardie et al, 2012)
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