| Objective:The abnormal metabolism and depostion of beta-amyloid protein(Aβ) is closely related with the pathogenesis of Alzheimer's disease(AD), which could cause the inflammatory response and toxic effects to synapse in AD. The change of synaptic structure can occur in the beginning of AD, earlier than extensive neuronal degeneration. Post synaptic density (PSD) is a vulnerable structure of synapse and plays an important role in synaptic plasticity. Triptolide(TP) is a kind of Chinese original drug with anti-inflammatory and immunosuppressive effects,but it has not been reported whether TP has a protective effect to synaptic plasticity in AD rat model. Therefore, we established an AD rat model by microinjecting aggregated Aβ1-40 into left hippocampus. Then we explored the protection mechanism of TP on synaptic plasticity by observing the expression changes of N-methyl-D-asparate receptor subunit 2B(NR2B) and postsynaptic density 95(PSD-95)in PSD of AD rat model and the effects of TP on them. These would provide experimental evidences for searching for effective traditional Chinese medicine.Method:1. Establish AD model by microinjecting aggregated Aβ1-40 into rat's left hippocampus, which simulates AD behaviors and pathological features, such as the impairment of learning and memory, glial cell proliferation and the neuronal damage. The rats injected with the same volume of saline were used as control.2. Morris water maze test was performed one week after the microinjection, by which we selected successfully established AD model rats, and divided them randomly into the AD model group and the TP-treated group. Rats in the TP-treated group were intraperitoneally injected with TP (0.4 mg/kg) every day, while the same volume of 4% propanediol was given to the AD model and control group. The intraperitoneal injection lasted for 15 days.3. Observe the histomorphology changes nearby the injection spot by nissl staining. Immunohistochemistry and RT-PCR were performed to investigate the PSD related proteins (NR2B and PSD-95) expression at protein and mRNA levels.4. Observe the ultrastructural changes of synapse in the hippocampus neurons by transmission electron microscope.Results:1. Nissl staining demonstrated that, after Aβ1-40 microinjection, the dorsal granule cell zone in dentate gyrus nearby the injection spot was incomplete, the neuron number decreased, and the glial reaction is general and severe. But in the TP-treated group, the neuron number only decreased slightly, and the glial cells was much less than in the AD model group, which suggest a protection effect of TP on hippocampus neurons from inflammatory injury induced by Aβ1-40 microinjection.2. Immunohistochemically, the number of the NR2B and PSD-95 staining positive cells of the TP-treated group were significantly more than those of the AD model group (P<0.01,P<0.05), and the average optical density of which were significantly increased (P<0.01), although still less or decreased compared to those of the control group (P<0.01). The immunohistochemical results demonstrate that TP could promote the NR2B and PSD-95 protein expressions probably by its anti-inflammatory effects.3. The RT-PCR results showed that the expression levels of NR2B and PSD-95 mRNA of the AD model group were significantly decreased, compare with the control group (P<0.01). The expression levels of NR2B and PSD-95 mRNA of the TP-treated group were higher than those of the AD model group (P<0.05), while still lower than those of the control group (P<0.05, P<0.01). The RT-PCR results demonstrate that TP could promote the NR2B and PSD-95 mRNA expressions probably by its anti-inflammatory effects.4. The results of transmission electron microscopy demonstrated that, in the synaptic number, synaptic vesicles and thickness of PSD in the hippocampus neuron, the model group showed significant decrease compared with the control group. However, after TP treatment, the synaptic number, synaptic vesicles and PSD thickness restored, which suggested a protection effect of TP on synapse ultrastructural changes coursed by Aβ1-40 microinjection. Conclusion:TP can protect the synaptic ultrastructure and promote PSD related proteins (NR2B and PSD-95) expressions both at protein and mRNA levels in the AD model rats, indicating that it may protect the synaptic plasticity of AD model rats.
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