老年痴呆实验动物模型 研制进展 Research Progress of Animal Models for Alzheimers Disease 张 斌 (Bin Zhang) M.D., Ph.D. Center for Neurodegenerative Disease Research, School of Medicine University of Pennsylvania Philadelphia, PA USA High Median Low Millions Year 美国 85岁以上的人口统计 US Population Demographics 85 Years of Age or Older: 1950-2050 Average life span in USA Male: 72.2 years old Females: 79.10 years old 美国老年痴呆发病率 AD Incidence In US AD afflicts 4 million Americans 100,000 die/year. AD occurs in men and women of all ethnic groups and at all socioeconomic levels. The US Census Bureau predicts that AD will affect 14 million Americans by 2050. 美国老年痴呆病人数量 (百万) AD Patients In USA 1980 2005 2050 老年痴呆神经病理 Neuropathology of AD 神经元纤维缠结(N FT ) Neurofibrillary Tangles Tau protein (细胞内) 老年斑(SP) Senile Plaques 淀粉样蛋白 -Amyloid (A) (细胞外) 理想的老年痴呆实验动物模型标志 Idea AD Animal Models 老年痴呆病人老年痴呆动物模型 _ 病理表现神经元纤维缠结 细胞内形成tau蛋白纤维 老年斑 细胞外形成 淀粉样蛋白纤维 生化特点 Tau蛋白超磷酸化 类似AD Ab, Tau蛋白降低溶解性类似AD 行为改变 记忆力下降或丧失, 行为测试显示 记忆力下降或丧失 _ 实验动物模型重复性, do not reduce the gene expressing level 实验动物模型实用性, drug screen, mechanism study 老年痴呆实验动物模型种类 Types of AD Animal Models 基因种类: 正常和变异的基因 Normal and mutant genes Tau 蛋白, Tau protein b淀粉样前蛋白 APP Presenin1, 2, PS1,2 载脂蛋白E, Apolipoprotein E (ApoE) 酶 b secratase Cyclin dependent kinase5 (CDK5) glycogen synthase kinase3 (GSK3) 动物种类: 非脊椎动物模型 果蝇模型 Drosophila model 线虫模型 Nematode Caenorhabditis elegans model 脊椎动物模型 小鼠模型 mouse model 大鼠模型 rat model Alzheimer-type neuropathology in transgenic mice overexpressing V717F b- amyloid precursor protein. Dora Games et al., Nature 373, 523-527 Tau transgenic C. elegans. Kraemer et al., PNAS, 100:9982-9985 Tau transgenic drosophila Jackson et al., Neuron 34:509-519 Gtz et al., Molecular Psychiatry 9, 664 老年痴呆实验动物模型种类(续) Types of AD Animal Models 基因数量 单基因模型 Single Tg model, Tau, APP, PS1 双基因模型 double Tg model, Tau+APP, APP+PS1 三基因模型 triple Tg model, Tau+APP+PS1 基因表达的细胞 神经元细胞 neuron 神经胶质细胞 Neuronal glial cell 少突胶质细胞 Oligodendrocyte 星形胶质细胞 Astrocyte Tau and APP double Tg mice. Lowis et al., Science 293:1487-1491 Makoto Higuchi, Bin Zhang, Mark S. Forman, Yasumasa Yoshiyama, John Q. Trojanowski, and Virginia M.-Y. Lee J. Neurosci., 25: 9434, 2005 Mark S. Forman,Devika Lal, Bin Zhang, Deepa V. Dabir, Eric Swanson, Virginia M.-Y. Lee,and John Q. Trojanowski The Journal of Neuroscience, 2005, 25:3539 1. Prepare your DNA + promoter 2. Transform fertilized eggs Harvest freshly fertilized eggs before the sperm head has become a pronucleus. Inject the male pronucleus with your DNA. When the pronuclei have fused to form the diploid zygote nucleus, allow the zygote to divide by mitosis to form a 2-cell embryo. 3. Implant the embryos in a pseudopregnant foster mother and proceed offsprings. -Over express protein 转基因动物模型 transgenic animal model. -基因敲除动物模型 gene knockout animal model - 基因敲入动物模型 gene knockin animal model -条件基因动物模型 conditional animal model 基因引入动物的方法 老年痴呆实验动物模型种类(续) Types of AD Animal Models 基因启动子种类 -人Prion 基因启动子 human PrP promoter (极微小的蛋白质颗粒，类似于病毒，但 不含核酸，被认为是绵羊痒病和某些神经系统变性疾病的传染介质) -鼠Prion 基因启动子 murine PrP promoter， -鼠CNP基因启动子 murine CNP olygodendrocyte promoter -鼠胶质原纤维酸性蛋白启动子 murine GFAP promoter -鼠钙调蛋白激酶IIa 启动子 Mouse CaMKIIa promotor (鼠产生的启动子， 钙调蛋白激酶IIa 驱动基 因表达在产后二周的前脑和海马部位) K. SantaCruz et al., Science, 309: 476- 481, 2005 I. 淀粉样蛋白连锁反应假说 The Amyloid Cascade Hypothesis By Dennis Selkoe, Harvard University II. 载脂蛋白对淀粉样蛋白的作用假说 Effects of ApoE on Amyloid-b Hypothesis By David M. Holtzman, Washington University III. 蛋白质错误折叠和轴突转运障碍假说 Tau Related Axonal Transport Dysfunction Hypothesis By John Trojanowski, University of Pennsylvania 2005 Update 老年痴呆发病机理研究 AD Pathogenesis Studies Dennis Selkoe, Nature. 360(6405):672-4 , Proc Natl Acad Sci U S A 91(25):11993-7. Cai XD et al., Science, 259 :514-6. Suzuki N et. al., Science, 264:1336-40 淀粉样蛋白连锁反应假说 The Amyloid Cascade Hypothesis II. 载脂蛋白对 A的作用假说 Effects of ApoE on Amyloid-b Hypothesis II. 载脂蛋白对 A的作用假说 (续) Effects of ApoE on Amyloid-b Hypothesis III. 蛋白质错误折叠和轴突转运障碍假说 Protein Misfolding and Axonal Transport Dysfunction Hypothesis Mechanisms Of Protein Misfolding And Brain Amyloidosis Forman M, Trojanowski, JQ, Lee VM-Y. Nat Med, 2004 a-synuclein tau vesicles Tau aggregation/hyperphosphorylation Decreased MT bound tau MT depolymerization Impaired axonal transport Axonal degeneration -appears to arrest cells in mitosis by stabilizing spindle MTs. -induce MT polymer mass in cell and “MT bundling”; -readily soluble in methanol, but limited soluble in water; PaxceedTM -A new MT Stabilizing anti-neoplastic agent by Angiotech Inc.; -In contrast to Taxol (Cremaphor formulation), PaxceedTM is a Micellar formulation that does not cause the inflammatory reactions; -Micellar paclitaxel is soluble in saline and once in the bloodstream, the micelles break down so t