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Protective Effects and Mechanisms of EGCG on Alzheimer’s Disease Mice

Author: HeMiao
Tutor: WeiMinJie
School: China Medical University
Course: Pharmacology
Keywords: (-) Epigallocatechin gallate catechin gallate D-galactose APP/PS1 transgenic mice Apoptosis Caspase-3 Amyloid precursor protein p75 neurotrophin receptor c-jun N-terminal kinase P53
CLC: R96
Type: PhD thesis
Year: 2008
Downloads: 395
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Abstract

The purpose of Alzheimer's disease (Alzheimer's disease, AD) is an occurrence in middle-aged neurodegenerative disease progressive cognitive impairment and memory ability. Its typical pathological change is the large number of β-amyloid-like peptides (β-Amyloid, Aβ) is the core of the senile plaques (senileplaques, the SP), the neurons in extracellular body appeared nervous original fiber tangles (neurofibrillary tangles in the nerve cells, NFT ) and the like. The pathogenesis of AD has not been fully elucidated, but the doctrine of free radical damage and apoptosis doctrine much attention. The formation of free radicals and oxidative stress and further induced neuronal apoptosis, is the basis of degradation and neurodegenerative AD functionality. p75 neurotrophin receptor the (P75 neurotrophin receptor, P75NTR) is a low affinity neurotrophin receptor the AD neuron apoptosis. p75NTR induced apoptosis signaling pathway is not yet very clear, but has been widely recognized that the activation of c-jun N-terminal kinase (c-jun N-terminal kinase, JNK)-mediated neuronal apoptosis key steps, and promote further by activating the expression of apoptotic proteins such as P53, Bad, promote or induce apoptosis. In addition, JNK can further activation of by inducible cytochrome C release Caspase-3 and leads to apoptosis. Research and the establishment of a reliable AD animal models proven AD etiology, pathogenesis and prevention of drug R \u0026 D have important significance. Far for many AD studies in animal models, wherein D-galactose (D-galactose, D-gal) to mice with chronic injection has become a common method, widely used in AD and other neurodegenerative diseases pharmacological studies. In addition, the transgenic model is a cause of model, Overexpression of human-derived mutations AD-related gene in transgenic mice, pathological and functional changes in part because of its clear cause, and can reflect AD, helps us understand the pathogenesis of AD becoming the overall model of AD. The prevention and treatment of AD anti-oxidant and anti-apoptotic drug research and development to become one of the important topics of today pharmacology research areas. (-) Epigallocatechin epigallocatechin gallate (epigallocatechin-3-gallate, EGCG) is a major green tea polyphenol, the study confirmed that green tea effective iron chelation, antioxidant, anti-inflammatory, anti-cancer and neuroprotective such as role play a major dependence of EGCG. Levites through the study of human into neuroblastoma SHSY5Y cells and rat fibroblasts tumor PC12 cells that EGCG can reduce the generation of the neurotoxic effects of Aβ. In this study, therefore, to be established by D-galactose subcutaneous injection of mouse models of AD and hereditary APP/PS1 transgenic mice, using Niss1 ??method study of by TUNEL staining, immunohistochemistry, Westernblot., D-galactose in mice cognitive behavior, pathological changes, antioxidant capacity, apoptosis, Aβ (1-40) amyloid precursor protein (Amyloid of precursor Protein, APP), promote apoptotic protein caspase-3 and apoptosis of P75-JNK-P53 related pathway, and 9-month-old APP/PS1 turn the indicators of change of the gene in mice, and further explore the neuroprotective effect of EGCG on two mouse models of AD and its mechanism. MATERIALS AND METHODS D-galactose induced AD model mice experiment: Take Kunming mice 96, were randomly divided into 6 groups (n = 16), the blank control group (Control), model group (D-gal ddH ₂ O), model group 2 (D-gal oil), the VE the positive control group (D-gal 280IU/kg-VE), EGCG low-dose treatment group (D-gal mg / kgEGCG)., EGCG high dose treatment group (D-gal 6 mg / kg EGCG). A day to two model group and VE positive control group, EGCG low-dose and high-dose treatment group mice 150 mg / kg subcutaneous injection of 3% of the D-galactose once blank control group of mice injected subcutaneously daily volume saline continuous injection six weeks. The third week to VE positive control group of EGCG low dose and high-dose group treated daily were 5.6% VE Press 280IU/kg gavage at 2 mg / kg orally 0.04% EGCG, press 6 mg / kg double distilled water gavage 0.12% EGCG once, blank control group and model group mice fed the same amount of time, the model group the same amount of soybean oil gavage once administered continuously for 4 weeks. The last week of the mice water maze, passive avoidance and locomotor activity behavior detection during continuous administration. Behavior after, quick to take the part of the mouse hippocampus into -80 ℃ cryopreservation, respectively of SOD, GSH-Px activity and MDA content determination and Western blot blot analysis of APP, caspase-3, P75, JNK2 and P53, etc. protein expression levels. Another part of the mouse brain perfusion fixation, respectively HE staining and immunohistochemical detection the Niss1 ??staining, TUNEL staining and Aβ (1-40), caspase-3, APP. APP/PS1 were collected from 20 of the 10 C57 mice and APP/PS1 mice, transgenic mice were randomly divided into three groups: the C57 the control group (Control), model group (APP/PS1), the experimental group (APP/PS1 2 mg / kgEGCG). EGCG daily to the experimental group APP/PS1 transgenic mice to 0.04% at 2 mg / kg orally once, to the control group and the model group mice fed the same amount of double distilled water intragastrically 4 weeks. The last two days of passive avoidance test mice behavior detection during continuous administration. Behavior after rapid take part the rat hippocampal into -80 ℃ cryopreservation, Western blot blot analysis of P75, JNK2 and P53 protein expression levels. Another part of the mouse brain perfusion fixation, HE staining and immunohistochemical detection of Aβ (1-40). Experimental results EGCG AD mice induced by D-galactose memory and judgment water maze, passive avoidance, locomotor activity behavior results show that D-galactose (150 mg / kg) subcutaneous injection of 6 weeks, mice showed learning apparent decline in memory and judgment, but the activity was not affected; EGCG (2 mg / kg and 6 mg / kg) and VE (280IU/kg), were significantly improved D-galactose induced AD model mice learning and memory impairment. EGCG D-galactose induced AD model mouse brain pathological changes in the impact of HE staining and Niss1 ??staining results showed that, six weeks after D-galactose (150mg/kg) subcutaneous injection, mice cerebral cortex and hippocampus nerve significantly reduce the number of cells, the cells loosely unclear boundaries Nissl bodies, necrotic neurons; EGCG (2 mg / kg and 6 mg / kg) and VE (280IU/kg), significantly reducing the D-galactose induced neuronal damage. 3, EGCG D-galactose induced AD model mice brain Aβ (1-40) protein expression Aβ (1-40) Immunohistochemical staining showed: D-galactose (150 mg / kg) subcutaneous injection after six weeks, the mouse cerebral cortex and hippocampus, Aβ (1-40) protein expression levels significantly increased; EGCG (2 mg / kg and 6mg/kg) and VE (280IU/kg), significantly inhibited mouse brain cortex and hippocampus Aβ (1-40) protein expression levels. 4, EGCG D-galactose induced AD mice hippocampus SOD, GSH-Px activity and MDA content enzymology results show that D-galactose (150 mg / kg) subcutaneous injection six weeks later, the mice hippocampus within the activities of SOD, GSH-Px enzyme activity significantly decreased MDA content increased significantly; EGCG (2 mg / kg and 6 mg / kg) and VE (280IU/kg), can significantly improve the mouse hippocampus SOD, GSH-Px activity, and significantly reduce the content of MDA. 5, EGCG brain regions of AD model mice induced by D-galactose neuronal apoptosis TUNEL staining results showed that AD model induced by D-galactose (1 50 mg / kg) of the rat cerebral cortex and hippocampus seen more apoptotic cells and apoptotic index with the blank control group increased significantly; EGCG (2 mg / kg and 6 mg / kg) and VE (280IU/kg), can significantly reduce the D-galactose induced neuronal cell apoptosis. AD model mice brain EGCG D-galactose-induced caspase-3 protein expression of Caspase-3 immunohistochemical staining and Western blot blot analysis showed that D-galactose induced mouse brain (150mg/kg) cortex and hippocampus, caspase-3 protein expression level was significantly higher; EGCG (2 mg / kg and 6 mg / kg) significantly inhibited the D-galactose induced mouse cerebral cortex and hippocampus, caspase-3 protein levels increased; VE (280IU/kg) significantly inhibited. EGCG AD model mice induced by D-galactose brain APP protein expression of APP immunohistochemistry and Western blot blot analysis showed that D-galactose (150 mg / kg) induced mouse cerebral cortex and hippocampus The area of ??the APP protein expression levels significantly higher; EGCG (2 mg / kg and 6mg/kg) and VE (280IU/kg), inhibited D-galactose induced mouse cerebral cortex and hippocampus of APP protein expression levels were significantly the increase. EGCG AD model mice induced by D-galactose hippocampus of P75NTR protein JNK2 and P53 protein expression Western blot blot analysis showed that D-galactose (150 mg / kg) significantly promoted the P75 protein within the mouse hippocampus shear increase the the nuclear transfer p75ICD expression level also significantly induced mouse hippocampus JNK2 and P53 protein expression levels increase; EGCG (2 mg / kg and 6 mg / kg) significantly inhibited D - the galactose induced by p75ICD, JNK2 and P53 protein expression levels increase; VE (280IU/kg) no significant inhibition. 9, EGCG APP/PS1 transferred gene the mice memory and ability to judge impact passive avoidance test results show APP/PS1 transgenic mice and C57 mice than there has been significant memory impairment; EGCG (2 mg / kg) significantly improved APP/PS1 transgenic mice memory impairment. 10, EGCG on the protein expression of Aβ (1-40) Immunohistochemical staining showed: APP/PS1 transgenic mice cerebral cortex and hippocampus of Aβ (1 APP/PS1 transgenic mouse brain Aβ (1-40) -40) protein expression levels of C57 mice was significantly higher; EGCG (2 mg / kg) can significantly reduce the the APP/PS1 turn transgenic mouse cerebral cortex and hippocampus of Aβ (1-40) protein expression levels. 11, EGCG APP/PS1 transfer the inside of P75NTR protein transgenic mouse hippocampus the JNK2 and P53 protein expression Western blot blot analysis showed the the APP/PS1 turn transgenic mouse hippocampus p75ICD JNK2 and p53 protein expression levels of C57 mice significantly higher ; EGCG (2 mg / kg) can significantly inhibit the APP/PS1 turn transgenic mouse hippocampus p75ICD, of JNK2 and P53 protein expression levels. Conclusion 1 EGCG can improve the D-galactose induced AD model mice and APP/PS1 transgenic mice learning and memory impairment. 2, EGCG can increase antioxidant enzyme activity in the hippocampus of AD model mice induced by D-galactose, to play an effective antioxidant effects. 3, EGCG protein levels by inhibiting the AD model mice induced by D-galactose in the cerebral cortex and hippocampus of APP and Aβ (1-40) of nerve cells play a protective role. 4, EGCG the transgenic mice hippocampus within p75ICD, JNK2 and p53 apoptotic pathway protein expression levels by inhibiting D-galactose induced AD model mice and APP/PS1 further inhibition of caspase-3 protein activity of nerve cells play an effective The anti-apoptotic effect.

Protective Effects and Mechanisms of EGCG on Alzheimer’s Disease Mice

Abstract

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