Protective Effects Of Icariin Preventive Treatment On Mitochondrial Damage In Alzheimer’s Disease

Alzheimer’s disease (AD) is a degenerative disease of the central nervous system, and its main clinical manifestations are memory decline and loss of cognitive function, especially ecmnesia. In recent years, the incidence of AD shows an increasing trend, but its pathogenesis is very complex, so far is not clear. It was suggested that mitochondrial oxidative stress and mitochondrial dysfunction may be involved in the pathological process of AD. Estrogen has a neuroprotective effect, and its neuroprotective effect is closly related with mitochondria. Epimedium has improved cognitive function, and in recent years it has been found a strong estrogenic activity. So we choose icariin (ICA) which is the main active ingredient of Epimedium and icaritin (ICT) which is the metabolites of ICA, using normal SD rats, SAMP 8 and ovariectomized SAMP8 (OVX SAMP8) mouse respectively to investigate the protective effects of preventive administration of icariin on mitochondrial damage in Alzheimer’s disease.Part 1 Effect of ICT on rat brain mitochondrial function in vitroObjective:Different concentrations of ICT were incubated with rat brain mitochondria in vitro to explore the effect of ICT on mitochondrial function. Methods: Mitochondria isolated from rat brain were incubated with different concentrations of ICT (0.01-10μmol/L) in 37 ℃ for 10min. Brain mitochondrial respiratory function, swelling of the membrane, the membrane potential, ATP content and ROS change were measured. Results:After incubated with different concentrations of ICT, the structure and function of rat brain mitochondria were changed. Brain mitochondrial respiratory function and ATP content were not obviously changed, only 0.01 μmol/L of ICT could increase P/O value of the NADH respiratory chain and 1μmol/L of ICT could reduce RCR of FADH2 respiratory chain. High concentrations of ICT could significantly damage mitochondrial structure, showing increased mitochondrial membrane swelling, decreased membrane potential and increased ROS levels.With increasing concentration of ICT, the degree of change are increasing. Conclusion: ICT could affect the structure and function of rat brain mitochondria in vitro. ICT could not significantly affect mitochondrial respiratory function and ATP content, which high concentration of ICT could significantly damage mitochondrial structure.Part 2 Effect of preventive administration of ICA on learning and memory abilities and brain mitochondrial damage in senescence-accelerated mouseprone8 (SAMP8)Objective:To investigate the effect of preventive administration of icariin on learning and memory abilities and brain mitochondrial function and brain mitochondrial oxidative stress in senescence-accelerated mouse prone8 (SAMP8). Methods:The 6-month-old SAMP8 mice were randomly divided into the SAMP8 model group, ICA groups (75,150 mg/kg), the positive Diethylstilbestrol (DES) group and estrogen receptor inhibitor ICI182780 combined with ICA high-dose (150 mg/kg) group, with 8 mice in each group.8 same month old SAMR1 mice were selected as the normal control group. After oral administration of ICA for 8 weeks, the uterus wet weight and serum E2 levels were measured to test estrogen-like effects of ICA. Morris water maze test and step-down passive test were used to investigate the effects of preventive administration of ICA on learning and memory abilities in SAMP8 mice. Cerebral cortex mitochondria were isolated to determine the effect of preventive administration of ICA on the brain mitochondrial function by detecting mitochondrial respiratory function, mitochondrial membrane swelling, mitochondrial membrane potential and ATP content. Effect of preventive administration of ICA on brain mitochondrial oxidative stress was detected by ROS level, MDA content, GSH content and CAT activity. Results:Preventive treatment of ICA could increase uterus coefficients. ICA could significantly improve the abilities of place navigation and space exploration of SAMP8 mice, and enhance their reflex ability in step-down passive test. ICA could improve brain mitochondrial respiration activity, reduce membrane swelling, increasing membrane potential and increase ATP content in brain mitochondria of SAMP8 mice. ICA could also reduce the level of ROS and MDA content in brain mitochondria of SAMP8 mice, but GSH content and CAT activity were not significantly increased. Compared with ICA 150mg/kg group, the uterus coefficients, learning and memory abilities, mitochondrial respiratory function and mitochondrial membrane potential levels of mice in ICA150mg/kg and estrogen receptor inhibitor ICI182780 co-administrated group were significantly decreased. However, brain mitochondria oxidative stress was not changed obviously. Conclusion:Preventive administration of ICA has estrogen-like effects, and ICA could significantly improve learning and memory abilities and brain mitochondrial function and brain mitochondrial oxidative stress in SAMP8 mice. The mechanism of ICA improving learning and memory abilities and brain mitochondrial function may be related to its estrogen-like effect; while the action on brain mitochondrial oxidative stress may be independent of estrogen receptor.Part 3 Effect of preventive administration of ICA on learning and memory abilities and brain mitochondrialdamage in OVX SAMP8Objective:To investigate the effect of preventive administration of icariin on learning and memory abilities and brain mitochondrial function and brain mitochondrial oxidative stress in OVX SAMP8. Methods:All the 6-month-old SAMP8 mice were removed their ovaries except mice in sham group. Mice were divided by the weight into the OVX SAMP8 model group, ICA groups (75,150 mg/kg) and the positive Diethylstilbestrol (DES) group, each group has 10 mice.10 same month old SAMR1 mice were selected as the normal control group. After oral administration of ICA for 8 weeks, the uterus wet weight and serum E2 levels were measured to test estrogen-like effects of ICA. Morris water maze test and step-down passive test were used to investigate the effects of preventive administration of ICA on learning and memory abilities in SAMP8 mice. Cerebral cortex mitochondria were isolated to determine the effect of preventive administration of ICA on the brain mitochondrial function by detecting mitochondrial respiratory function, mitochondrial membrane swelling, mitochondrial membrane potential, ATP content. Effect of preventive administration of ICA on mitochondrial oxidative stress were detected by ROS level, MDA content, GSH content, SOD activity and CAT activity. Results:Preventive treatment of ICA could increase uterus coefficients and E2 levels. ICA could significantly improve the abilities of place navigation and space exploration of OVX SAMP8 mice, and enhance their reflex ability in step-down passive test. ICA could improve brain mitochondrial respiration activity, reduce membrane swelling, increase membrane potential and ATP content in brain mitochondria of OVX SAMP8 mice. ICA could also reduce the level of ROS and MDA content, increase SOD activity in brain mitochondria of OVX SAMP8 mice, but GSH content and CAT activity were not significantly increased. Conclusion:Preventive administration of ICA could improve endogenous estrogen of OVX SAMP8 mice, which means ICA has estrogen-like effects. Meanwhile, ICA could significantly improve learning and memory abilities and brain mitochondrial function and brain mitochondrial oxidative stress in OVX SAMP8 mice.