Mechanism Study Of The Protective Effects Of Icariin On Learning And Memory Ability Via Inhibiting Inflammatory Response In AD Mice

Alzheimer's disease(AD)is the highest incidence of neurodegenerative diseases of central nervous system.AD is characterized by the progressive memory loss and cognitive impairment.Inflammation has been proved to participate in the pathologic process of chronic neurodegenerative diseases as an independent factor.As the most abundant component of the Traditional Chinese medicine herba epimedii,icariin(ICA)has been shown a strong anti-inflammatory and neuroprotective functions in a variety of inflammatory associated diseases models both in vivo and in vitro.It has been confirmed that ICA can restrain the release of proinflammatory factors of microglia to exert its anti-inflammatory and neuroprotective effects.But the target protein and the related signaling pathways are still unclear.As we all know,ICA has oestrogen-like activity.The newly discovered membrane estrogen receptor-G protein coupled estrogen receptor(GPER)has been demonstrated to mediate the growth of neuronal synapses in rat hippocampus.Binding with ligand,GPER will active growth factor receptor,adenylate cyclase,phospholipase C as well as calcium ion signaling pathways via G protein.Our previous studies have confirmed that ICT,the derivative of ICA,could protect against MPP+-induced damage in dopaminergic MES23.5 cells via the insulin-like growth factor-1 receptor(IGF-1R)signaling pathway.MEK/ERK and PI3K/Akt signaling pathways are involved in the neuroprotective effects of ICA against MPTP-induced dopaminergic neuronal damage.So we put forward whether ICA could suppress the activation of microglia and alleviate inflammation in hippocampus via GPER and IGF-1R signal pathways.In the present experiment,the AD inflammation model was established by intracerebroventricularly injection of lipopolysaccharide(LPS)in ICR mice.Morris water maze,real time PCR,western blot and immunohistochemistry were used to detect the mechanisms of ICA against AD inflammation.The experimental results are as follows:1.According to the results of Morris water maze,contrast to the control group,mice in the LPS group took longer latency and less quadrant time of training and number of platform crossings(P<0.01,P<0.001);According to the results of real time PCR,gene expressions of i NOS,COX-2,IL-1? and TNF-? in hippocampus were significantly increased(P<0.001);According to the results of western blot,the protein expressions of the limited enzyme of proinflammatory cytokines i NOS and COX-2 in hippocampus were significantly increased(P<0.001).Different dosages of ICA(5mg/kg,10mg/kg,20mg/kg)could inhibit LPS-induced inflammation in varying degree,and the 10mg/kg ICA had the strongest effect;it could significantly reduce latency,enhance the quadrant time of training and number of platform crossings(P<0.01,P<0.001),decrease the gene expressions of i NOS,COX-2,IL-1?and TNF-?(P<0.01,P<0.001)as well as the protein expressions of the limited enzyme of proinflammatory cytokines i NOS and COX-2 in hippocampus(P<0.05,P<0.001).2.The improving effect of ICA on LPS-induced learning and memory damage could be blocked by GPER antagonist G15(0.2?g/1ml)in AD model mouse.Compared with the ICA group,mice in ICA,LPS and G15 co-treatment group took longer latency and less quadrant time of training and number of platform crossings(P<0.001);gene expressions of TNF-?,IL-1?,i NOS and COX-2 in AD model mouse were significantly increased(P<0.05,P<0.01);the protein expressions of the limited enzyme of proinflammatory cytokines i NOS and COX-2 in hippocampus were also increased(P<0.01,P<0.001);There was no significant difference between G15+LPS group and LPS group.3.LPS could significantly increase the phosphorylation levels of MAPKs(ERK,JNK,p38)(P<0.01,P<0.001).ICA could protect against the protein phosphorylation induced by LPS(P<0.05,P<0.01).These effects could be blocked by G15(P<0.05,P<0.01).4.LPS could significantly increase the phosphorylation level of IkB(P<0.001).ICA could protect against the protein phosphorylation induced by LPS(P<0.001).These effects could be blocked by G15(P<0.001).5.The improving effect of ICA on LPS-induced learning and memory damage could be partly blocked by IGF-1R antagonist JB-1 in AD model mouse.Compared with the ICA group,mice in ICA,LPS and JB-1 co-treatment group took longer latency and less quadrant time of training(P<0.05),but the number of platform crossings had no significant change(P>0.05);gene expressions of COX-2 and TNF-a were increased(P<0.05),while i NOS and IL-1? were slightly but not significantly increased(P>0.05);the protein expressions of the limited enzyme of proinflammatory cytokines i NOS and COX-2 in hippocampus were also slightly but not significantly increased(P>0.05);There was no significant difference between LPS+JB-1 treatment group and LPS group.6.The results of Immunohistochemisty showed that ICA treatment remarkably suppressed microglial activation induced by LPS.Co-treatment with G15 or JB-1abolished the inhibition of ICA on LPS-induced microglial over-activation,while the blocking effect of G15 was more significant.The above data indicate that ICA has inhibitory effect on LPS-induced inflammation in AD model mouse.The mechanism might be related to the activation of GPER and IGF-1R signaling pathways.This study provides a new target for neurodegenerative disease therapy.