NR_045396/MicroRNA761/FADD axis regulates necroptosis and survival of retinal ganglion cells.

The involvement of necroptosis and the underlying mechanism in retinal ganglion cell (RGC) death is not fully understood. We aim to determine whether the NR_045396/miRNA761/Fas-associated protein with death domain (FADD) axis participates in the regulation of necroptosis in RGCs. A mouse model of optic nerve crush was employed for in vivo experiments. Apoptosis and necrosis were assessed by TUNEL and Propidium iodide (PI) exclusion. We found that the necrotic rate increased in a time-dependent manner in RGCs following optic nerve injury, whereas apoptosis peaked at 7 days following nerve damage. Immunohistochemistry revealed that the expression levels of key markers of necroptosis, pRIP3 and pMLKL, were upregulated, whereas FADD expression was reduced in RGCs at 14 days after optic nerve injury. Enforced expression of FADD in RGCs by an AAV vector attenuated necrotic response and promoted RGC survival. A dual-luciferase reporter gene assay showed that miR761 directly regulated FADD expression. Intraocular application of AAV2 expressing sequences complementary to miR761 binding site (AAV2-miR761 sponge) enhanced FADD expression and regulated RGC necrosis and survival. Moreover, the long non-coding RNA (lncRNA) NR_045396 binds directly to miR761 and modulates the necrotic program of RGCs. Thus, we demonstrate the anti-necroptosis and neuroprotective effects of the NR_045396/miR761/FADD axis.