P3‐306: Major gene expression changes in the brain of presymptomatic anti‐NGF AD11 mice

not available. P3-307 MORRONISIDE PREVENTS APOPTOSIS IN HUMAN NEUROBLASTOMA CELLS BY H2O2INDUCED Wen Wang, Department of Pharmacology, Xuan-Wu Hospital of Capital Medical University, Beijing, China. Contact e-mail: lzwwang@yahoo.com.cn Background: Morroniside belongs to an extensive group of natural iridorid glycosides and extracted from the Chinese herb, Coruns officinalis Sieb. Methods: In the present study, using human neuroblastoma SH-SY5Y cells, we have investigated the protective effects of this compound on odifications in endogenous reduced glutathione (GSH), intracellular oxygen species (ROS) and apoptotic death on H2O2-mediated cytoxicity. Results: Incubation of cells with morroniside led to a significant dosedependent elevation of cellular GSH accompanied by a marked protection against H2O2-mediated toxicity. Morroniside at 1-100 lM inhibited the formation of ROS and the activation of caspase-3 and 9, and the upregulation of Bcl-2, whereas no significant change occurred in Bax levels. The results indicated that the anti-oxidative and anti-apoptotic properties render this natural compound potentially protective against H2O2-induced cytotoxicity. Conclusions: This study suggested that intracellular GSH appeared to be an important factor in morroniside-mediated cytoprotection against H2O2-toxicity in SH-SY5Y cells. P3-308 EARLY EXPRESSION OF THE FAS-SIGNALING ADAPTOR PROTEIN FADD IS LINKED WITH HYPERPHOSPHORYLATED TAU PROTEIN AT THE PRECLINICAL STAGE OF ALZHEIMER’S DISEASE Chuang-Kuo Wu, Changiz Geula, Edward Stopa, Z. Mao, X. Wang, Northwestern University, Chicago, IL, USA; Brown University, Providence, RI, USA; Emory University, Atlanta, GA, USA. Contact e-mail: chuangwu@northwestern.edu Background: The cause of cell death in Alzheimer’s disease (AD) is still under vigorous investigation. Although studies have demonstrated that beta-amyloid peptides can induce apoptosis, mechanisms of hyperphosphorylated-tau (P-tau) associated neurodegeneration remain elusive. Previously, we discovered that a Fas-signaling adaptor protein, FADD, is linked with P-tau proteins within tangle-containing cholinergic neurons in the advanced stage of Alzheimer brains. To test the hypothesis that tau hyperphosphorylation precedes the induction of FADD which could lead to cell death, we investigated the expression of the FADD protein and P-tau in the control brain and brains with minimal AD pathology, in comparison with brains with severe AD pathology. Methods: In first experiment, we used paraffin-embeded brain tissues. In second experiment, we employed fresh-frozen brain tissues for immunohistochemistry and western blotting analysis. The expression of P-tau was demonstrated by utilization of antibodies labeling site S396/404 (PHF-1) and site S202 (AT8). Using double-labeling immunohistochemistry, we studied the colocalization of FADD and P-tau and compared between brains with severe AD (Braak stage V-VI) and normal-control brains/brains with mild AD (Braak stage I-II). In both groups, the same three areas were examined: amygdale, entorhinal cortex and hippocampus. A quantitative study was undertaken. Results: Abundant FADD-immunoreactive structures, including neurofibrillary tangles and dystrophic neurites, were already observed in the entorhinal cortex, amygdala and hippocampus of the preclinical stage of Alzheimer brains (Braak stage I-II). By double-labeling methods, FADDimmunoreactivity was proved to be co-localized with P-tau positive tangles and dystrophic neurites and thus already present in brains with mild Alzheimer pathology. The expression of P-tau, as predicted in quantitative studies, was prominent in severe AD brains. Surprisingly, brains with mild AD pathology already displayed the similar level of FADD protein to that of severe AD brains. Conclusions: Abundant FADD expression was already observed even in the brains with low AD pathology (Braak stage I-II). This finding suggests that, at the preclinical stage, the Fas-signaling death system has been activated to be involved with P-tau associated neurodegeneration. The intracellular adaptor protein FADD can be the chaperon protein which promotes the abnormal phosphorylation of tau proteins in the very early phase of AD. P3-309 OVEREXPRESSION OF HYPERPHOSPHORYLATED TAU ANTAGONIZES A -POTENTIATED APOPTOSIS WITH PRESERVATION OF MITOCHONDRIAL ELEMENTS Jun Yin, Jian-Zhi Wang, Pathophysiology Department, Tongji Medical College, Hua-Zhong University of Science and Technology, Wuhan, China. Contact e-mail: yinjuntongji@yahoo.com.cn Background: We have recently reported that phosphorylation of tau antagonizes cell apoptosis (Li et al., PNAS, 3591-6, 2007). And A is a recognized apoptotic inducer in the brains of Alzheimer’s disease (AD). We further studied whether tau could antagonize A -potentiated apoptosis and the underlying mechanisms. Methods: N2a single cell clones with stable expression of APPswe (N2a/APPswe) or the vector (N2a/vector) were generated by selection with 250 g/mL G418. To study the effects of tau on the apoptosis, N2a/APPswe cells were transiently expressed tau or its vector plasmids and the HEK293 cells were transiently co-expressed the APPswe and tau or the vector plasmids. The cell apoptosis was induced by 250 M H2O2. Results: N2a/APPswe cells bearing high level of A were more vulnerable than the controls to H2O2-induced cell death. Concomitantly, elevation of Bax and cytosolic cytochrome-c and decrease of Bcl-2, as well as increase of nuclear fragmentation/activation of caspase-3 and poly (ADP-ribose) polymerase (PARP) were observed. Expression of the hyperphosphorylated tau in N2a/APPswe attenuated the A -potentiated apoptosis, demonstrated by reduced nuclear fragmentation and inhibition of caspase-3/PARP, and elevated Bcl-2 and suppressed Bax and cytosolic release of cytochrome-c. The anti-apoptotic effect of tau was also observed by transient co-expression of tau and APPswe in HEK293 cells, with a elevation of -catenin. Conclusions: We conclude that A renders the cells more vulnerable to apoptosis whereas expression of the hyperphosphorylated tau can attenuate the A -potentiated cell apoptosis with preservation of mitochondrial elements. P3-310 SEI-1, A CDK4 REGULATOR, IS INDUCED AND IS REQUIRED FOR NEURON DEGENERATION IN RESPONSE TO TROPHIC FACTOR DEPRIVATION AND -AMYLOID EXPOSURE Subhas C. Biswas, Lloyd A. Greene, Columbia University, New York, NY, USA. Contact e-mail: scb34@columbia.edu Background: A growing body of evidence indicates that aberrant activation of a cell cycle pathway is necessary for degeneration of neurons in cellular models of Alzheimer’s Disease (AD). Cell cycle proteins are also elevated in affected neurons of human patients with AD. However, how the cell cycle pathway is initiated in AD or AD models is unclear. Among the first steps in this pathway is activation of cyclin-dependent kinase 4 (Cdk4). In proliferating cells, the protein SEI-1 regulates Cdk4 activity and promotes DNA synthesis. Post-mitotic neurons remain in the G0 state in presence of trophic support such as nerve growth factor (NGF). It has been shown that lack of trophic support leads to degeneration of neurons. The loss of synaptic connections and axonal degeneration that occur in AD T612 Poster Presentations P3: