Celastrol supports survival of retinal ganglion cells injured by optic nerve crush

• We evaluated the effect of celastrol on RGC survival after optic nerve crush.
• Systemic and intravitreal injection of celastrol protect RGCs from axonal injury.
• Celastrol-mediated RGC protection is associated with downregulation of TNF-alpha.

The present study evaluates the effect of celastrol on the survival of retinal ganglion cells (RGCs) injured by optic nerve crush (ONC). Celastrol, a quinine methide triterpene extracted from the perennial vine Tripterygium wilfordii (Celastraceae), has been identified as a potential neuroprotective candidate in a comprehensive drug screen against various neurodegenerative diseases. Two weeks after ONC, the average density of remaining RGCs in retinas of animals treated with daily intraperitoneal (i.p.) injections of celastrol (1 mg/kg) was approximately 1332 cells/mm2, or 40.8% of the Celastrol/Control group. In retinas of the Vehicle/ONC group about 381 RGCs/mm2 were counted, which is 9.6% of the total number of RGCs in the DMSO/Control group. This corresponds to approximately a 250% increase in RGC survival mediated by celastrol treatment compared to Vehicle/ONC group. Furthermore, the average RGC number in retinas of ONC animals treated with a single intravitreal injection of 1 mg/kg or 5 mg/kg of celastrol was increased by approximately 80% (760 RGCs/ mm2) and 78% (753 RGCs/mm2), respectively, compared to Vehicle/ONC controls (422 cells/mm2). Injection of 0.2 mg/kg of celastrol had no significant effect on cell survival, with the average number of RGCs being 514 cells/mm2 in celastrol-treated animals versus 422 cells/mm2 in controls. The expression levels of Hsp70, Hsf1, Hsf2, HO-1 and TNF-alpha in the retina were analyzed to evaluate the roles of these proteins in the celastrol-mediated protection of injured RGCs. No statistically significant change in HO-1, Hsf1 and Hsp70 levels was seen in animals with ONC. An approximately 2 fold increase in Hsf2 level was observed in celastrol-treated animals with or without injury. Hsf2 level was also increased 1.8 fold in DMSO-treated animals with ONC injury compared to DMSO-treated animals with no injury suggesting that Hsf2 induction has an injury-induced component. Expression of TNF-alpha in retinas of celastrol-treated uninjured and ONC animals was reduced by approximately 2 and 1.5 fold compared to vehicle treated animals, respectively. The observed results suggest that mechanisms underlying celastrol׳s RGC protective effect are associated with inhibition of TNF-alpha-mediated cell death.