The Ketone Metabolite Attenuates Oxidative Stress And Inflammation After Spinal Cord Injury

Spinal cord injury(SCI)is a devastating neuro-destructing event causing motor and sensory deficits,with significant emotional and social disturbances,neuropathic pain,and cardiovascular complications.The pathophysiology of SCI can be classified into a primary mechanical injury and a secondary process orchestrated by diverse pathophysiological mechanisms,including breakdown of the blood-spinal cord barrier,neuroinflammation,oxidative stress,neuronal injury,ischemic dysfunction,and other processes.Ketogenic diet(KD)has been used to treat drug-resistant epilepsy,and has been shown various beneficial to many neurodegenerative diseases.Recent study demonstrated that POHB could suppress oxidative stress by inhibion of-class I histone deacetylases(HDAC)and nucleotide-binding domain and leucine-rich repeat protein 3(NLRP3)inflammasome.The ketone metabolite,?-hydroxybutyrate(?OHB),is reported neuroprotective after spinal cord injury(SCI)in rats,but the underlying mechanism remains unknown.The present study aims to investigate effects of ?OHB on suppression of oxidative stress through inhibition of class I histone deacetylases(HDACs)and inflammation through inhibition of NLRP3 inflammasome in in-vivo and in-vitro models.Rats were fed with ketogenic diet(KD)or standard diet(SD).A C5 hemi-contusion injury was applied to these animals on the 14th day of experiment,and spinal cord samples were harvested on the 1st,3rd and 7th day after SCI,respectively.Behavioral and electrophysiological assessments were conducted continuously until 12 weeks after SCI.KD significantly increased blood ketone levels and promoted forelimb functional and electrophysiological recovery.KD reduced oxidative stress markers and ROS products,down-regulated the expression of Nox2 and Nox4,and up-regulated the expression of Foxo3a,MnSOD and catalase after SCI.KD suppressed microglia activation and overproduction of inflammatory cytokines,down-regulated the expression of NLRP3,ASC and caspasel p20.KD also transformed the microglias from M1 to M2.The in-vitro study,performed on PC 12 cells,indicated that POHB inhibited H2O2-induced ROS production,decreased Nox2 and Nox4 protein levels,and up-regulated Foxo3a,MnSOD and catalase levels in a dose-dependent manner,which was consistent with the in vivo results.The ketone metabolite ?OHB inhibited HDAC1,HDAC2 and HDAC3 activity,but not HDAC8 in SCI rats and PC 12 cells.Depletion of HD AC 1 or HDAC2 with siRNA attenuated H2O2-induced ROS production and protein carbonylation,elevated Foxo3a protein levels,meanwhile reduced Nox2 and Nox4 protein expression in PC 12 cells.The in-vitro study,performed on BV2 cells,indicated that ?OHB inhibited LPS-induced inflammatory response,decreased NLRP3 protein levels.Inhibiation of NLRP3 by MCC950 or siRNA could transformed the microglias from Ml to M2.Our results indicate that the ketone metabolite ?OHB attenuates oxidative stress in spinal cord injury by inhibition of class I histone deacetylases,and selected suppression of HDAC1 or HDAC2 regulates Foxo3a,Nox2 and Nox4 expression.We also demonstrate that ketone metabolite?OHB suppress inflammation through inhibiation of NLRP3 inflammasome,and transformed the microglias from M1 to M2.Thus,the ketone metabolite ?OHB may be a novel promising therapeutic agent for spinal cord injury.