【摘要】：目的：建立缺氧缺糖神經細胞損傷模型，探討EPO發(fā)揮神經保護作用的可能機制，希望為EPO在神經系統疾病的臨床應用提供理論依據。 方法：將出生后24h內C57BL/6小鼠的大腦皮質神經元原代培養(yǎng)7-10d后，神經元特異性烯醇化酶（Neuron specific enolase, NSE）免疫組化染色方法鑒定神經元的純度；用含連二亞硫酸鈉的無糖Earle’s液模擬大腦缺血缺氧性損傷，建立缺氧缺糖神經細胞損傷模型。分組：1、正常組；2、模型組（缺氧缺糖神經細胞損傷模型）；3、EPO組(EPO組干預的缺氧缺糖神經細胞損傷模型）。MTT檢測細胞存活率、FITC-Annexin V/PI熒光染色方法檢測細胞凋亡率、測定LDH漏出率，觀察EPO對缺氧缺糖損傷神經元的保護作用。 結果：1、成功原代培養(yǎng)神經細胞，經NSE免疫組化方法鑒定神經元純度大于90%；2、建立缺氧缺糖神經細胞損傷模型，模型組中神經細胞凋亡率為（15.2±0.37），LDH漏出率為（20.4±0.33），細胞存活率為（54.1±0.51），與正常組相比差異有統計學意義（P0.01）；3、與模型組相比，EPO組的細胞凋亡率、LDH漏出率低于模型組，細胞存活率高于模型組（P0.01）。 結論：原代培養(yǎng)小鼠神經細胞，成功構建缺氧缺糖神經細胞損傷模型。EPO能夠減輕缺氧缺糖對神經元的損傷，，該保護作用可能是通過抑制神經細胞凋亡途徑介導完成。
[Abstract]:Objective: to establish a model of hypoxia-glucose deficiency nerve cell injury and to explore the possible mechanism of neuroprotective effect of EPO in order to provide a theoretical basis for the clinical application of EPO in nervous system diseases. Methods: after primary culture of cerebral cortical neurons of C57BL/6 mice within 24 hours after birth for 7-10 days, the purity of neurons was identified by immunohistochemical staining with neuron-specific enolase (Neuron specific enolase, NSE). A model of hypoglycemic neuronal injury was established by simulating cerebral ischemia and hypoxia injury with Earle's solution containing sodium bisulfite. Groups: 1, normal group, 2, model group (hypoxia and glucose deficiency nerve cell injury model); (3). MTT, FITC-Annexin V/PI fluorescence staining and LDH leakage were used to detect cell survival rate, apoptosis rate and LDH leakage rate in EPO group (hypoxia / glucose deficient nerve cell injury model). To observe the protective effect of EPO on neurons injured by hypoxia and glucose deficiency. Results: 1. The primary cultured neurons were successfully cultured, and the purity of neurons was higher than 90 by NSE immunohistochemical method. 2. In the model group, the apoptosis rate was (15.2 鹵0.37), LDH leakage rate was (20.4 鹵0.33), and the cell survival rate was (54.1 鹵0.51). Compared with the normal group, the difference was statistically significant (P0.01). 3Compared with the model group, the apoptosis rate and LDH leakage rate in EPO group were lower than those in the model group, and the cell survival rate was higher than that in the model group (P0.01). Conclusion: EPO can attenuate the neuronal damage induced by hypoxia and glucose deficiency, which may be mediated by inhibiting neuronal apoptosis.
【學位授予單位】：遵義醫(yī)學院
【學位級別】：碩士
【學位授予年份】：2014
【分類號】：R741

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