| Alzheimer's disease (AD) is a neurodegeneration disorder that significantly impairs the memory and congnitive functions of the brain. The morbidity of AD in the elder is only lower than that of angiocardiopathy and cancer. Pathological changes in AD are characterized by the formation of senile plaques and neuroflbibrillary tangles as well as extensive neurons loss. Amyloid beta protein (AP), the major ingredient of senile plaques, plays a critical role in the development of AD. Although the concrete mechanisms of Ap-induced neurotoxicity are unclear, the loss of Ca2+ homeostasis, the generation of oxidative stress and the enlargement of excitatory toxicity consider being involved. The disturbance of intracellular Ca2+ homeostasis may affect energy in processes and material metabolism, and arouses a succession of pathphysiologic reaction. It is believed to be the final common pathway to neuron death by which various agents manifest their neurotoxicity. Therefore, efforts to investigate the pathway of changes in Ca2+- homeostasis-induced by AP may provide a way to protect against Ap neurotoxicity and a novel therapeutic strategy approaches for AD. Basic fibroblast growth factor (bFGF) is a pluripotent cytokine that canenhance the survival and proliferation of neurons and protect neurons against insults relevant to ischemia injury and brain trauma. There are many experiments focus on neuroprotection of bFGF domestic, but there is no studies related with AD. The neuroprotective effects of bFGF on cultured hippocamal neurons with AP will be studied in our experiments. We expect to provide some evidences for using bFGF to treat AD.Objective: (1)To investigate the neurotoxicity effects of Ap and the protective effects of bFGF on damage neurons induced by A1-40 or H2O2 in cultured hippocampal neurons. (2)To study the possible pathway of the intracellular free calcium concentration changed by A1-40.Methods: (l)The cultured newborn rat hippocamal neurons were treated with aggregated A1-40 with different concentration for 24h. MTT assay and LSCM binding Fluo-3/AM were used to detect the survival rate and the intracellular free calcium concentration of neurons respectively. (2)Neurons were pretreated with bFGF for 24h and then treated with A1-40 or H2O2. By using the same marker, bFGF as a protective agent, we observed the protective effect of bFGF on damage induced by Ap1-40 or H2O2. (3)A real time method was applied to measure the level of free calcium concentration in neurons with LSCM, we observed the influence of the L-type VDCC blocker nimodipine and NMDA receptor blocker MK-801 on calcium influx changed by Ap1-40.Result: (1)Morphological study show that neurons treated by A1-40 for 24h were shrinkage of the cell soma, neurite fragmentation, and eventually lysis. The OD values were 0.302+0.030, 0.254+0.026, 0.133+0.009 respectively in MTT experiment and the level of [Ca2+]i were 2.53+0.76. 3.17+0.72, 6.97+0.74 respectively in hippocampal neurons treated with 0.1 , 1,10mol/L A1-40 for 24h.The survival rate was decreased and fluorescence intensity was increased following the increased Ap1-40 concentration. There was significantly difference between higher dosage group and control group (P<0.05), but low dosage group had no difference (P<0.05). (2)Neurons were pretreated with 20ng/ML bFGF and then treated with Ap1-40 or H2O2 for 24h, the OD value were 0.319+0.011, 0.314+0.027, 0.300+0.019 or 0.378+0.068, 0.380+0.04, 0.355+0.051, the fluorescence intensity were 2.26+0.68, 2.38+0.76, 2.53+0.61 or 2.49+0.65, 2.44+0.57, 2.58+0.32 respectively. There was no difference between all exposed group and control group (P>0.05). (3)Ap1-40 could not increases [Ca2+]i in free-calcium D-Hank's solutions, but had a side action in cultured medium. Nimodipine partly protect [Ca2+]j increasing induced by Ap1-40, but MK-801 had no inhibition.Conclusion: (1)The morphology was changed and the survival rate was reduced when hippocampal neurons were treated with aggregated A1-40, and this action was coincidence wit...
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