BDNF/TrkB And SIRT1are Involved In The Antagonistic Role Of Hydrogen Sulfide In Homocysteine-induced Neuronal Injury In The Hippocampus Of Rats

[Background and Objective]Homocysteine (Hcy)-elicited neuronal cell death has been implicated in thedevelopment or pathology of Alzhermer’s disease (AD) patients. Hydrogensulfide (H2S) has recently been recognized as an important endogenousneuroprotectant, including antioxidant, anti-apoptosis actions. We havepreviously demonstrated the protective effect of H2S against Hcy-inducedneurotoxicity. However, the mechanisms responsible for this beneficial role of H2Sare unclear.Brain-derived neurotrophic factor (BDNF) is secreted by the neurons. It playsan important role in neuronal differentiation, survival, and development. Silentinformation regulator (SIRT1), one member of the family of Sirtuins, is expressed inthe brain, cerebellum and hippocampus and plays an important role in cellmetabolic, proliferation, and aging. Based on these, we will explore whether themechanism underlying the protection of H2S against Hcy-induced neurotoxicity isinvolved in BDNF and SIRT1.Therefore, our study used SD rats intracerebroventricularly injected with Hcyas an animal model of Hcy-induced neurotoxicity to explore whether H2Santagonises Hcy-induced neuronal injury in hippocampus and whetherBDNF/TrkB and SIRT1mediates the antagonistic role of H2S in homocysteine-induced neuronal injury in hippocampus.[Methods]Neuronal apoptosis in hippocampus was analysed byTerminal-deoxynucleotidyl transferase-mediated dUTP nick-end labeling (Tunel)staining. Expressions of Glucose regulated protein78(GRP78), C/EBPhomologous protein (CHOP) and cleaved caspase-12in hippocampus wasmeasured by Western blot. Malondialdehyde (MDA) levels in the hippocampusof rats were detected by Elisa.[Results]1. Homocysteine induces neuronal injury in hippocampus.Tunel staining indicated that Hcy (0.6,2.0μmol/d×7d, icv) induced apopticneurons in the CA1area in hippocampus of rats, which revealed that Hcyinduced neuronal apoptosis in the hippocampus of rats.Hcy (0.2,0.6,2.0μmol/d×7d, icv) concentration-dependently increases theexpressions of GRP78, CHOP and cleaved caspase-12in the hippocampus of rats,which indicated that Hcy induced ER stress in the hippocampus of rats.Results of Elisa indicated that Hcy (0.2,0.6μmol/d×7d, icv) increases thelevel of MDA in the hippocampus of rats, which demonstrated that Hcy causesoxidative stress in the hippocampus of rats.2. Hydrogen sulfide antagonizes homocysteine-induced neuronal injury inhippocampus.Tunel staining showed that NaHS (100μmol/kg/d×9d, ip) markedlyameliorated the apoptotic neurons in hippocampus induced by Hcy (0.6μmol/d×7d, icv), which indicated that H2S resists apoptosis of neurons inducedby Hcy.NaHS (100μmol/kg/d×9d, ip) attenuated Hcy (0.6μmol/d×7d)-induced increase in the expressions of GRP78, CHOP and cleaved caspase-12inhippocampus of rats, which indicated that H2S inhibitis Hcy-inducedhippocampal ER stress.The result of Elisa showed that NaHS (100μmol/kg/d×9d, ip) significantlyantagonized the increasing level of MDA induced by Hcy (0.6μmol/d×7d, icv) inhippocampus of rats, which suggested that H2S antagonizes Hcy-inducedhippocampal oxidative stress.3. K252a, the specific inhibitor of BDNF/TrkB, reverses the antagonistic role ofHydrogen sulfide in homocysteine-induced neuronal injury in hippocampus.Tunel staining showed that K252a (1μg/d×9d, icv), the specific inhibitor ofTrkB (the receptor of BDNF), can obviously reverse the antagonistic role of NaHS(100μmol/kg/d×9d, ip) in Hcy (0.6μmol/d×7d, icv)-increased numbers ofapoptotic neurons in hippocampus, which revealed that BDNF/TrkB pathway isinvolved in the inhibitory role of H2S in Hcy-induced neuronal apoptosis in thehippocmpus of rats.Western blotting results indicated that K252a (1μg/d×9d, icv) can obviouslyreverse the antagonistic role of NaHS (100μmol/kg/d×9d, ip) in Hcy (0.6μmol/d×7d, icv)-increased expressions of GRP78, CHOP and cleavedcaspase-12in the hippocampus of rats. These data demonstrated that BDNF/TrkBpathway is involved in the inhibitory role of H2S in Hcy-induced hippocampal ERstress.The result of Elisa showed that K252a (1μg/d×9d, icv) can obviously reversethe antagonistic role of NaHS (100μmol/kg/d×9d, ip) in Hcy (0.6μmol/d×7d,icv)-induced increase in the level of MDA, which suggested that BDNF/TrkBpathway is involved in the antagonistic role of H2S in Hcy-induced hippocampaloxidative stress. 4. Sirtinol, the specific inhibitor of SIRT1, reverses the antagonistic role ofHydrogen sulfide in homocysteine-induced neuronal injury of hippocampus.Tunel staining showed that Sirtinol (10nmol/d×9d, icv), the specific inhibitorof SIRT1, can obviously reverse the antagonistic role of NaHS (100μmol/kg/d×9d,ip) in Hcy (0.6μmol/d×7d, icv)-induced increase in the number of apoptoticneurons in hippocampus, which suggested that SIRT1is involved in the inhibitoryrole of H2S in Hcy-induced neuronal apoptosis in the hippocmpus of rats.Western blotting results indicated that Sirtinol (10nmol/d×9d, icv) canobviously reverse the antagonistic role of NaHS (100μmol/kg/d×9d, ip) in Hcy(0.6μmol/d×7d, icv)-induced increase in the expressions of GRP78, CHOP andcleaved caspase-12in the hippocampus of rats, which demonstrated that SIRT1is involved in the inhibitory role of H2S in Hcy-induced hippocampal ER stress.The result of Elisa showed that Sirtinol (10nmol/d×9d, icv) can obviouslyreverse the antagonistic role of NaHS (100μmol/kg/d×9d, ip) in Hcy (0.6μmol/d×7d, icv)-induced increase in the level of MDA, which suggested thatSIRT1is involved in the antagonistic role of H2S in Hcy-induced hippocampaloxidative stress.[Conclusion]1. H2S prevents Hcy-induced neuronal injury in the hippocampus of rats.2. BDNF/TrkB pathway and SIRT1are involved in the antagonistic role of H2S inHcy-induced neuronal injury in hippocampus.