Therapeutic Effect Of Koumine On Experimental Alzheimer’s Disease And Its Possible Mechanism

Objective: To investigate the effect of koumine on the dominant learning and memory impairment manifestation of Alzheimer’s disease(AD)and to observe the effect of koumine on characeristic senile plaques and neurofibrillary tangle pathology upon okadaic acid(OA)-induced AD-like model in rats,in order to propose the novel therapeutic effect of koumine on AD and to provide the experimental proofs for the potential clinical application of koumine as a hopeful drug in the prevention and treatment of AD.To explore the relationship between the therapeutic effect of koumine on AD and the expression of amyloid precursor protein(APP),the activation of microglia and astrocytes,the expression of TNF-α and IL-1β,undertaken to elucidate the possible mechanism for the therapeutic effect of koumine on AD and provide the theoretical foundation on the appliance of koumine and make further recognition of pathogenetic mechanisms of AD.Methods:(1)AD-like symptoms were imitated by multiple intracerebroventricular injection of OA in rats.Koumine hydrochlorate(0.056,0.28 or 1.4 mg/kg)was administered intragastricly,once a day for ten consecutive days starting from the day next to the last day of OA injection.Evaluate the effect of koumine on the learning and memory ability of rats using place navigation test and spatial probe test by Morris water maze.Rat brain was harversted after the behavioral study was finished.Senile plaques and neurofibrillary tangle pathology in rat hippocampus was observed by Bielschowsky stain,in order to investigate the effect of koumine on AD-like characteristic pathological changes.(2)The expression of hippocampal APP was detected by western blot to analyze the relationship between the therapeutic effect of koumine on AD and the expression of hippocampal APP.(3)Positive immunoreactivity and the expression of Iba-1(a hallmark for microglia)and GFAP(a hallmark for astrocyte)in hippocampus was observed by immunohistochemistry and western blot respectively,in order to clarify the relationship beween the therapeutic effect of koumine on AD and the activation of microglia and astrocyte in hippocampus.(4)The expression of pro-inflammatory cytokine TNF-α and IL-1β was detected by western blot to elucidate the relationship beween the therapeutic effect of koumine on AD and the expression of TNF-α and IL-1β in hippocampus.Results:(1)Compared with control group,OA significantly increased the escape latency in the place navigation test and decreased the platform-site crossover times in the spatial test.This result suggests that AD-like symptoms are successfully imitated by multiple intracerebroventricular injection of OA in rats.Compared with model group,koumine(0.056-1.4 mg/kg)significantly decreased the scape latency in the place navigation test and increased the platform-site crossover times in the spatial test,which is indicated that koumine may alleviate the impairment of learning and memory in the experimental AD rats.Hippocampus senile plaques and neurofibrillary tangles were observed in OA rats while less in koumine-treated rats by Bielschowsky stain,which is implied that koumine may ameliorate the characteristic pathology of AD.(2)Western blot data shew that the expression of hippocampal APP in OA rats was higher than that in control rats,while the expression of hippocampal APP in koumine-treated rats was lower than that in OA rats.The result was negatively correlated with the result of spatial probe test,which is suggested that the therapeutic effect of koumine on AD may be related with the inhibition of the expression of APP in hippocampus.(3)It has been shown that positive immunoreactivity and the expression of Iba-1 and GFAP was increased in OA rats compared with control rats,while positive immunoreactivity of Iba-1 and GFAP was decreased in koumine-treated rats compared with OA rats.The results was negatively correlated with the result of spatial probe test,which is suggested that the therapeutic effect of koumine on AD may be related with the activation of Iba-1 and GFAP in hippocampus.(4)Western blot results showed that the level of hippocampal TNF-α and IL-1β was higher in OA rats than that in control rats,while the level of hippocampal TNF-α and IL-1β was lower in koumine-treated rats than that in OA rats.The result was negatively correlated with the result of spatial probe test,which is suggested that the therapeutic effect of koumine on AD may be related with the inhibition of the expression of TNF-α and IL-1β in hippocampus.Conclusion: This is the first report about the effect of koumine on AD.1.Koumine can significantly alleviate the impairment of memory and the characteristic pathological changes in experimental AD rats.2.The therapeutic effect of koumine on AD might be related with the inhibition the expression of hippocampal APP,the alleviation of the activation of microglia and astroyte in hippocampus,and the reduction of the expression of proinflammatory cytokine TNF-α and IL-β in hippocampus.