Protective Role Of NAD~+ On Cognitive Function In Aged Rats With Chronic Cerebral Hypoperfusion

Background and AimChronic cerebral hypoperfusion,a pathological state,exists in the process of Alzheimer’s disease,vascular dementia and other diseases,which can lead to cognitive impairment in varying degrees.The pathogenesis and interventions of CCH-related cognitive impairment have been the focus of the researches.In this study,we constructed a SD rat model with chronic cerebral hypoperfusion and cognitive impairment,and then gave nicotinamide adenine dinucleotide intervention to explore the protective mechanism of NAD⁺on cognitive impairment in model rats,which may be related to NAD⁺regulation of mitophagy.Methods1.Five aged SD rats were randomly selected from twenty ones for cerebral blood flow monitoring alone（no further for follow-up tests）.2.The left 15 aged SD rats were randomly divided into 3 groups（n=5）:sham-operated group（namely sham group）;chronic cerebral hypoperfusion group（namely CCH group）;CCH model and NAD⁺intervention group（namely NAD⁺group）.For CCH group,a modified two-step bilateral common carotid artery occlusion（two-step BCCAO）method was used to establish animal model of chronic cerebral hypoperfusion.For NAD⁺group,after CCH model was finished,rats were injectedβ-NAD solution（250μg/g/d,PH=7）for 4 weeks.3.Morris water maze（MWM）was employed to test spatial reference memory.4.Laser speckle contrast imaging（LSCI）was used to monitor cerebral blood fluid（CBF）changes.5.NAD⁺/NADH kit was used to detect NAD⁺/NADH ratio in hippocampus.6.The relative expression of APP、TOMM20、LC3B、P62、SIRT1、PGC-1α、PINK1、Parkin in hippocampus were detected by Western blots.Results1.The escape latency of CCH group compared with sham group（51.35±5.58vs33.70±7.66,54.10±8.89vs23.15±11.03,54.25±9.25vs20.10±17.01,P<0.05）on the 2^rd,3^thh and 5^thh was prolonged and the difference was statistical significance.The escape latency of NAD⁺group was more shortened than that of CCH group（18.95±14.96vs54.10±8.89,P<0.01;18.50±13.65vs53.00±10.22,P<0.01;22.10±19.69vs54.25±9.25,P<0.05）on the 3^rd,4^thh and 5^thh and the difference was significant.2.Times of crossing the platform in CCH group was less than that in sham group（0.40±0.55vs 2.20±0.84,P<0.05）,while that in NAD⁺group was more than that in CCH group（2.60±1.10vs 0.40±0.55,P<0.05）.3.The percentage of target quadrant swimming time in CCH group was significantly lower than that in sham group（39.33±3.83 vs 25.67±2.79,P<0.01）,while that in NAD⁺group was higher than that in CCH group（40±6.97 vs25.67±2.79,P<0.05）.4.Immediately after BCCAO,the CBF decreased to baseline CBF（73.53±11.26）%,P<0.05;4 weeks after BCCAO,the CBF decreased to baseline CBF（71.81±9.16）%,P<0.01.5.There was no significant difference in the total content of NAD⁺and NADH between CCH group and sham group,but the ratio of NAD⁺/NADH in CCH group was lower than that in sham group（P<0.05）;the total content of NAD⁺and NADH and the ratio of NAD⁺/NADH in NAD⁺group were higher than that in CCH group（all P<0.05）.6.The ratio of NAD⁺/NADH was positively correlated with the times of crossing the platform（P<0.001）,and the ratio of NAD⁺/NADH was positively correlated with the percentage of swimming time in the target quadrant（P<0.01）.7.The expression levels of LC3B,PINK1,Parkin,APP（all P<0.05）and P62,TOMM20（all P<0.01）in CCH group were higher than those in sham group;the expression levels of APP（P<0.01）and TOMM20,LC3B,P62（all P<0.05）in NAD⁺group were lower than those in CCH group;the expression levels of SIRT1（P<0.05）and PGC-1α（P<0.001）in NAD⁺group were higher than those in CCH group.Conclusion1.The spatial learning and memory of CCH aged rats decreased significantly,and improved significantly after supplement of NAD⁺precursors.2.The cognitive protective effect of NAD⁺may be related to the inhibition of over-activated mitophagy and the balance of autophagy degradation function.3.NAD⁺can regulate PINK1/Parkin dependent mitophagy pathway through SIRT1-PGC-1αpathway,and play a protective role in cognitive impairment of CCH rats.