| ObjectTo observe the effect of lactoferrin on the cognitive function of aging mice,and explore the relationship between lactoferrin improvement aging mice cognitive function and oxidative stress,chronic inflammation and mitochondrial function.Methods16-month-old C57/BL6 J mice were randomly divided into Aged group,Aged+ LF group,and 2-month-old C57/BL6 J mice were used as young control group(Young group).According to previous studies,the daily gavage of lactoferrin administered was 2 g/kg/d,the other two groups were given normal saline for 3months.Three months later,Morris water maze experiment was applied to explore spatial learning and memory ability of different groups.Changes of synapses in hippocampal CA1 region were observed by transmission electron microscopy.Western-blot was used to detect the proteins expression levels of Synaptophysin(SYN)and Postsynaptic density 95(PSD 95)to evaluate synaptic function.The Superoxide dismutase(SOD),Malondialdehyde(MDA)and Reactive oxygen species(ROS)activities of hippocampus tissue were measured to evaluate the oxidative stress response by assays kits.Enzyme linked immunosorbent assay(ELISA)was used to measure the content of interleukin-1(IL-1β),interleukin-6(IL-6)and Tumor necrosis factor(TNF-α)in serum of mice.In order to assess mitochondrial morphological changes,we have observed the mitochondrial morphology of hippocampus tissues by transmission electron microscopy and quantify analysis by biomedical stereology.Real-time fluorescence quantitative PCR and Western-blot were used to measure the mitochondrial Dynamic fusion related proteins: Optic alrophy 1(Opa1),Mitofusin 2(Mfn2),and mitochondrial fission protein: Dynamic related protein 1(Drp1)genes and protein expression levels,to assess mitochondrial dynamics.Results1.Lactoferrin improved the cognitive function of aging mice.Morris water maze experiment results showed that the escape latency of Aged group was significantly longer than the Young group,the frequency of crossing the platform and the proportion of using spatial search strategy were lower.While the escape latency of Aged + LF group was significantly shorter than Aged group,the frequency of crossing the platform and the proportion of using spatial search strategy were increased.It suggested that spatial learning and memory ability of mice in the drug administration group were enhanced than the Aged group.The transmission electron microscopy showed that synaptic ultrastructure of Aged group was obscurer,postsynaptic density was thinner,and quantity was decreased than the Young group.While the synaptic ultrastructure of Aged + LF group was clearer,postsynaptic density was thickened and the quantity was increased than Aged group.Meanwhile,western blot results showed that the expression of SYN and PSD 95 in the Aged + LF group were significantly increased than Aged group.2.Lactoferrin have improved aging mice mitochondrial function.Oxidative stress testing results showed that Aged group mice brain tissues ROS,MDA levels were increased and the activity of SOD was decreased than Young group.While the Aged + LF group levels of ROS and MDA were decreased,the activity of SOD was up-regulated.ELISA results showed that the content of Aged group mice serum inflammatory factors(IL-1β、IL-6、TNF-α)were increased than Young group.After lactoferrin treatment,the content of Aged + LF group mice serum inflammatory factors(IL-1β、IL-6、TNF-α)were decreased than Aged group.The transmission electron microscopy showed that mitochondrial structure of Aged group was swollen and mitochondrial cristae was obscurer than the Young group.Biomedical stereology quantitative analysis displayed,under the premise of none significant difference in mitochondrial number and volume densities,the specific surface area of mitochondria in Aged group was decreased than Young group.While the mitochondrial structure of Aged + LF group was relatively clearer,mitochondrial cristae was clearer than Aged group,and the specific surface area of mitochondria in Aged + LF group was increased,mitochondrial swelling was alleviated.RT-PCR detection results showed that Aged group mitochondria fission genes(Drp1)mRNA expression was increased and fusion genes(Opa1,Mfn2)mRNA expression were decreased compared with Young group.While Aged+ LF group Drp1 mRNA expression was decreased and Opa1,Mfn2 mRNA expression were increased.The expression levels of Drp1,Mfn2 and Opa1 proteins were detected by western blot consistent with RT-PCR.ConclusionLactoferrin intervention improved the aging mice cognitive function,and its mechanism may be related to reduced oxidative stress,chronic inflammatory,and then improve mitochondrial morphology and function in the aging mice brain. |
