FGF21-mediated Protective Effect Of Methionine Restriction On Cognitive Impairments In Aged Mice

With the increasing of aging population over the world,age-related neurodegenerative diseases have become a serious threat to human health.Age-related neuronal damage and decreased synaptic plasticity have severely affected memory,language,cognition,and behavior.Recent years,researchers found that dietary restriction can promote health aging and has become a hot topic in the field of food nutrition.Calorie restriction and intermittent fasting have been shown to prolong life span and reduce the risk of aging related diseases in a variety of animal models,such as cerebrovascular disease,type 2 diabetes,cancer.The latest research shows CR and IF have potential intervention on the development of AD.Methionine restriction(MR)is one of the dietary restrictions widely reported in recent years,which can improve metabolism and prolong life span of aging animals.In this study,MR diet was conducted to investigate the improvement of age-related cognitive impairments and synaptic plasticity damages,and further reveal the nutritional response of FGF21 and its potential molecular mechanism of regulating oxidative stress in the nervous system.(1)In order to investigate the effect of MR on cognitive function of aging mice,2-month-old,12-month-old and 15-month-old C57BL/6 male mice were fed with different dietary patterns for 3 months: i)normal methionine diet(CD,Met 0.86%);ii)methionine restricted diet(MR,Met 0.17%).Behavioral experiments were tested after the diet intervention and tissue samples were collected for histological tests.In the open field test,Y maze test,and Morris water maze test,the cognitive behavior of aging mice decreased significantly compared with young mice.MR improved the autonomous activity ability of15-month-old aging mice,and significantly improved the decline of cognitive memory ability of 12-month-old and 15-month-old mice.The results of HE staining showed that MR improved the nuclear pyknosis and deepening of staining induced by aging.The effect of MR on the synaptic structure of aging mice was evaluated by transmission electron microscope.The results showed that MR could significantly increase the length of PSD.Also,MR improved the damages of synaptic structure induced and significantly increase the level of PSD95 in aging mice.Therefore,MR can significantly improve the age-related cognitive impairment and neurological damage.(2)In order to investigate the effect of MR on the function of hippocampal neurons in aging mice,the expression of genes in hippocampus was analyzed by RNA-seq.Using DEG analysis,we found that there were significant changes in neural related pathways such as Alzheimer’s disease pathway,energy metabolism related pathways such as oxidative phosphorylation pathway in the hippocampus of aging mice.Using GSEA gene enrichment,we found that MR had significant changed myelin function related genes and FGFR activation related genes.Moreover,we detected the levels of FGF21 in serum,liver.and brain of mice by ELISA.MR significantly activated the circulating FGF21 level in aging mice.The RNA-seq data was integrated analysis with the mouse characterizations by using DAIBLO method.It was found that there was a high correlation between the behavioral performance,FGF21 level and neural related genes in hippocampus.Therefore,FGF21 signal may play a key role in improving cognitive impairment in aging mice.(3)In order to reveal the mechanism of FGF21 nutritional response signal on improving cognitive and memory function of aging mice,2-month-old and 15-month-old C57BL/6 male mice were used.sh FGF21 was carried by adeno-associated virus and injected via tail vein to silence the expression of FGF21.CD or MR intervention were performed as before.In the open field test,new object recognition test,Y maze test,and Barnes maze test,MR improved the cognitive function of aging mice.Knockdown of FGF21 by i.v.injection of adeno-associated virus abolished the neuroprotective effects of MR in aged mice.Further,t SH-SY5 Y nerve cells were treated with r FGF21,the results showed that r FGF21 activated Nrf2 signaling pathway,and increase the expression of antioxidant enzymes HO-1 and NQO1.Therefore,FGF21 signal activation is required for MR improving cognitive function of aging mice.(4)In order to investigate the sex difference of MR in improving cognitive function of aging mice,male and female 15-month-old C57BL/6 mice were used.The results showed that there was no significant sex difference in body weight and food intake.The results of behavioral experiments showed that MR could improve the working memory ability and spatial memory ability of male and female aging mice.The ultrastructure of hippocampus was observed by TEM.It was found that there was no significant sex difference in the improvement of postsynaptic density.There was no significant difference in the number of mitochondria in hippocampus between male and female mice.Therefore,there was no significant sex difference in the activation of FGF21 nutrition response signal.In conclusion,MR can activate Nrf2 antioxidant signaling pathway by increasing the expression of FGF21,improve oxidative stress,protect synaptic plasticity,and improve cognitive impairment and nerve cell damage caused by aging.This study provides a theoretical basis for dietary patterns to improve neurodegenerative diseases caused by aging,and provides a new idea for studying the mechanism of dietary restriction patterns to improve cognitive dysfunction.