Effect Of Saponins Of Panax Japonicus On Brain Tissue In Natural Aging Rats Involvment Of Oxidative Stress And Autophagy Pathways

Objective To investigate the effects of saponins from Panax japonicus (SPJ) on braintissue in aging rats involvement of oxidative stress and autophagy pathways and study theeffects of SPJ on oxidative stress-induced injuries of SH-SY5Y cells so as to further definethe mechanism of SPJ against oxidative stress in aging rats.Methods Experiment one: the SPF male SD rats were randomly divided into5groups:3months group (3M group),9months group (9M group),15months group (15M group),24months aging group (24M group), SPJ low, medium and high dose groups,10rats ineach group. SPJ was irrigated into the animals with a dosage of10mg/kg,30mg/kg,60mg/kg from the18th month to the24th month. Intragastric administration stopped twodays a week. After6months, the morphological changes of hippocampus neurons in CA1,CA3and DG regions were observed by HE staining; the hippocampus synapticmorphology were observed under a transmission electron microscope; the activity ofSOD, GSH-Px, Na+/K+-ATPase, Ca2+-ATPase, Ca2+/Mg2+-ATPase and the content of MDAin cortex were measured by biochemical method; p21, Bcl-2, Bax, Sirt1, PGC-1α, Foxo3a,LC3II and Beclin1protein expressions in hippocampus and cortex were dectected bywestern blot. Experiment two: normal SH-SY5Y neurons were divided into normal controlgroup, H2O2(600μmol/L) group and H2O2+SPJ (0.1μg/mL,1μg/mL,5μg/mL,20μg/mL) group. Cells were pretreated with SPJ for12h followed by H2O2treatment for12h. After the indicated treatment, cell viability was detected by MTT; intracellular reactiveoxygen species was evaluated by DCFA-DA probe; LDH release quantity, the activity ofSOD, the MDA content, the ROS content and the mitochondrial membrane potential weremeasured by biochemical method; hochest staining analysed the apoptosis of SH-SY5Ycells and Bcl-2, Bax, Sirt1, PGC-1α, Foxo3a, LC3II and Beclin1protein expressions weredectected by western blot.Results Experiment one: Compared with young rats of3M group, hippocampus neurons inCA1, CA3and DG regions arranged in disorder with irregular shapes and hyperchromaticnuclei; the edge of nucleus is not clear and there are a lot of lipofuscin deposition inhippocampal neurons and p21protein expression increased significantly and the ratio ofBcl-2/Bax decreased significantly in hippocampus and cortex of aging rat of24M group.Compared with aging rat of24M group, SPJ each dose group significantly improved themorphological changes of CA1, CA3and DG neurons of hippocampus and the ultrastructure of hippocampal neurons; reduced p21protein expression and increased theratio of Bcl-2/Bax in hippocampus and cortex; obviously increased the activity of SOD andGSH-Px and decreased the content of MDA in cortex; effectively reversed the activity ofNa+/K+-ATP, Ca2+-ATP and Ca2+/Mg2+-ATPase and promote the Sirt1, PGC-1α, Foxo3a,LC3II and Beclin1protein expressions in hippocampus and cortex. Experiment two:Compared with control group, cell viability decreased significantly; the release of LDH,the content of MDA and the intracellular ROS was significantly increased and the activityof SOD and MMP decreased significantly. Compared with H2O2group, SPJ increasedH2O2-induced cell viability loss in SH-SY5Y cells; reduced the cell supernatant of LDHrelease quantity, the MDA content, the ROS content and the mitochondrial membranepotential and increased the activity of SOD (P <0.05, P <0.01); hoechst33342stainingobservation found that SPJ each dose group improved apoptosis obviously and Westernblot analysis found that SPJ each dose group raised the ratio of the Bcl-2/Bax andpromoted Sirt1, PGC-1α, Foxo3a, LC3II and Beclin1protein expressions.Conclusion SPJ exerted protective effects on aging rats and H2O2injured SH-SY5Y cellsand its mechanism may be related to the regulation of SPJ on oxidative stress andautophagy levels.