Amphiregulin and Growth Differentiation Factor 11 Promote Retinal Ganglion Cell Survival In Vivo through Activation of Smad3

Although adult neurons in the mammalian central nervous system do not spontaneously regenerate after injury, growing evidence indicates that genetically manipulating them can increase their ability to regenerate their axons. However, as genetic manipulation is not clinically feasible, current research continues to investigate pharmacological approaches to transiently enhance the intrinsic ability of adult CNS neurons to survive and regenerate their axons. Amphiregulin is a unique epidermal growth factor receptor ligand that has been shown to be crucial in liver regeneration, and accumulating evidence suggests that AREG signaling can promote both survival and axon regeneration of neurons. Growth differentiation factor 11 is a member of the transforming growth factor β superfamily, and it has been shown to exert rejuvenation effects in the aged brain and promote neuronal survival in the CNS. However, it is unknown whether AREG and GDF11 can induce neuronal survival and axon regeneration in the visual system. Hence, the current study investigated: 1) the developmental and post-injury expression pattern of these two ligands and their respective receptors, EGFR and activin-like kinase 5, 2) the potential neuroprotective and regenerative effects of these two ligands on retinal ganglion cells by using optic nerve crush model, and 3) the molecular mechanisms mediating the neuroprotective and regenerative effects of AREG and GDF11 on RGCs. Based on the western blot and immunohistochemical data, both AREG and GDF11 are consistently expressed throughout the retina development, but their receptors are only upregulated during the early retina development. Furthermore, AREG expression is significantly reduced in the adult retina 7 days after ONC whereas GDF11 expression remains unchanged after ONC. Interestingly, while EGFR expression is only upregulated 3 days after ONC, ALK5 expression is consistently upregulated throughout the post-injury time course. A single intravitreal injection of AREG or GDF11 immediately after ONC promoted significant RGC survival by activating Smad2/3 pathway. Because both ligands were not able to promote RGC axon regeneration, activation of Smad2/3 pathway may promote RGC survival but suppress RGC axon regeneration. Overall, the findings indicate that both AREG and GDF11 hold therapeutic potential for both neurodegenerative diseases and retinal degenerative diseases.