Study Of Cultured Hippocampal Neurons' Death Manner Induced By Amyloid-beta Protein In Vitro And The Anti-apoptotic Action Of NGF

Alzheimer's disease is a neurodegenerative disease that is characterized by progressive hypomnesia, cognition dysfunction and dystrophy. There are some pathologic characters in AD, for example, senile plaques, neurofibrillary tangles and loss of neuron's number. Amyloid beta protein is the core of SP and the biochemical foundation of NFT. Aβ has a direct toxicity to neuronal cells, but it is unclear that the percentage of apoptosis or necrosis induced by Ap. NGF is a neurotrophic factor, and its protective effect of NGF on the neuronal neurons degeneration occurred on AD is disputed.Objective: To study the toxicity of Aβ to hippocampal neurons. To investigate the protective effect of NGF on hippocampal neuron apoptopsis induced by Aβ.Methods:1. Neurons were dissociated from the hippocampal of newborn SD rats, and cultured with adding cytosine arabinoside, cytosine arabinosideand nerve growth factor. We observed neural morphologic changes. The survival rate of neurons cultured with different time was measured by invert microscope and MTT staining, and the rate of purification neurons by immunocytochemcal staining.2. The toxicity of pβ1-40 to neurons was observed in different concentrations and with different time-periods in vitro. Neuron death was analyzed by electron microscope DNA agarose gel electrophoresis and fluorescence staining.3. The apoptotic hippocampal euron was observed by transmission electron microscope, DNA electrophoresis and TUNEL staining in Aβ group and NGF group.Result:1.The growth of neurons added cytosine arabinoside and nerve growth factor was well, the purity and survival rate of neurons were high. The neurons purification was 98?.2% and survival ratye was 94.52 when they were exposed to NGF at 12 days.2. (1) The apoptosis and necrosis rates of neurons treated with 5 10 20mol/L Aβ1-40 were significantly higher than that of the control group after 12h. (2)At 6h, 12h and 24h incultion followed 10umol/L Aβ1-40 administration in the culturing medium, neural apoptosis and necrosis were significantly higher than that of the control group. (Neurons apoptotic rate was 30.5+6.51 and necrosis rate was 8.16?.62 when they were exposed to 10umol/LAβ1-40 for 12 hours).3. NGF can protect hippocampal neurons from toxicity induced by Aβ1-40, and reduced the rate of apoptosis, which was dose-dependented.(neurons apoptotic rate was 15.3+3.26 or 30.5+6.51 when they were treated with 10umol/L Aβ1-40 and NGF or 10umol/L Aβ1-40 respectively).Conclusion :1 NGF has a nutritional and supportive effect on neuronal cells.2 A certain concentrations of Aβ1-40 showed cytotoxic effect on cultured neurons and induced the neuron apoptosis or necrosis in a dose and time dependent manners.3 NGF can protect neuronal apoptosis from Aβ1-40.