The Effects Of Carnosine On Akt/mTOR Pathway And Autophagy In Hippocampus Of Rats With Vascular Cognitive Impairment

ObjectiveThe aim of this study is to investigate the impact of Carnosine(CAR)on the cognitive abilities of rats suffering from Vascular cognitive impairment(VCI).Additionally,we aim to identify the potential mechanisms involved by analyzing oxidative stress,Akt/m TOR pathway,and autophagy.This research will contribute to a better understanding of the effects of CAR and provide a theoretical foundation for its application.MethodsIn vivo experiments,54 SPF-grade male SD rats were selected,and after one week of adaptive feeding,the rats were randomly divided into 5 groups,namely the sham group,the model group and the CAR100,300 and 900 groups,including 10 in the Sham group,13 in the Model group,10 in the CAR100 group,11 in the CAR300 group and 10 in the CAR900 group.The VCI model was constructed by the modified two-vessel occlusion(2-VO)method,in which the CAR(100,300,900)group was given 100,300 and 900mg/kg carnosine daily gavage from 3 days before model preparation to 20 days after surgery.The Morris water maze experiment measures the spatial learning and memory ability of rats.Nissl staining to observe the degree of damage of neurons in the CA1 region of hippocampal tissue;The kit method NBT method detected SOD,and the Thiobarbituric acid method determined the MDA content;Western blotting method detected the expression of Akt,m TOR,p-Akt,p-m TOR,Beclin-1,LC3 B and p62 proteins in hippocampal tissues.Immunofluorescence colocalization staining to detect autophagy expression in hippocampal neurons.In vitro experiment,SH-SY5 Y cells were cultured and randomly divided into Control group(Control),model group(OGD/R),OGD/R+CAR(0.3,0.6,1.2 mmol/L)groups.Cell survival rate was detected by MTT method.The protein expressions of Akt,m TOR,p-Akt,p-m TOR,Beclin-1,LC3 B and p62 in each group were detected by Western blotting.ResultsThe results in vivo experiments showed that: 1.The escape latency of rats in the Model group was significantly longer than that in the Sham group(P<0.01),and the time of staying in the target quadrant was shorter than that in the sham group(P<0.01).At the same time,Nissl staining in the hippocampal CA1 region showed that the density and number of nerve cells were significantly reduced,the shape of the cells was changed(shrunk),the arrangement was loose and disordered,and the staining was not uniform.Compared with the Model group,after CAR treatment,the escape latency was shortened(P<0.01),the target quadrant residence time was longer(P<0.01 or P<0.05),the morphology of nerve cells in the CAR group was more complete,and the effect was more obvious with the increase of dose in the CAR group.2.The activity of SOD in the hippocampus of the Model group was significantly lower than that of the Sham group(P<0.01),and the content of MDA showed an upward trend(P<0.01),suggesting that the antioxidant capacity of the hippocampus of the VCI rats was weakened.The SOD activity in the CAR group was significantly higher than that in the Model group(P<0.01),and the MDA content was significantly lower than that in the model group(P<0.01).3.Western blotting showed no significant change in total Akt and total m TOR for each group.Compared with the Sham group,the p-Akt/Akt,pm TOR/m TOR ratios and p62 protein expression in the Model treatment group were decreased(P<0.01),and the Beclin-1/β-actin and LC3BII/LC3 BI ratio were increased(P<0.01).Compared with the Model group,the p-Akt/Akt,p-m TOR/m TOR ratios and p62 protein expression were increased(P<0.01 or P<0.05),and the Beclin-1/β-actin and LC3BII/LC3 BI ratio were decreased(P<0.01 or P<0.05)in the CAR treatment group.4.Immunofluorescence showed that the autophagy degree of Model group was enhanced compared with Sham group.Compared with Model group,autophagy degree of CAR group was reduced.In vitro experiment: 1.Compared with the control group,the cell viability of OGD/R group was significantly decreased(P<0.01).Compared with the OGD/R group,the cell viability of the OGD/R+CAR(0.3,0.6,1.2mmol/L)groups was significantly increased(P<0.01).2.There were no significant changes in total Akt and total m TOR for each group.Compared with the control group,the p-Akt/Akt,pm TOR/m TOR ratios and p62 protein expression in the OGD/R group were decreased(P<0.01),and the Beclin-1/β-actin and LC3BII/LC3 BI ratio were increased(P<0.01).Compared with the OGD/R group,the p-Akt/Akt,pm TOR/m TOR ratios and p62 protein expression were increased(P<0.01 or P<0.05),and the Beclin-1/β-actin and LC3BII/LC3 BI ratio were decreased(P<0.01 or P<0.05)in the OGD/R+CAR(0.3,0.6,1.2mmol/L)group.ConclusionsThe administration of CAR has been shown to enhance the cognitive abilities of VCI rats and mitigate the extent of nerve cell damage.These effects are likely attributed to the inhibition of oxidative stress,activation of the Akt/m TOR pathway in neurons,and suppression of nerve cell autophagy.