摘要
目的在乳鼠背根神经节神经元原代纯化培养方法的基础上,采用硝普钠(SNP)制备背根神经节神经元损伤模型。方法取乳鼠背根神经节,加入5-氟-2'-脱氧尿苷(5-FUDR)以纯化细胞,MEM-F12培养基培养,用神经元特异性烯醇化酶(NSE)免疫组织化学染色鉴定神经元。神经元培养至第八天分别加入不同浓度SNP(500,50,5μmol·L-1)制备背根神经节神经元损伤模型,用台盼蓝染色进行细胞计数,MTT法测定培养神经元活力。结果培养的脊髓背根神经节神经元生长状态正常,纯化培养纯度高于90%,可存活两周以上。培养的背根神经节神经元加入SNP作用后,随着浓度增加,神经元死亡率逐渐升高。结论采用SNP制备的脊髓背根神经节神经元损伤模型稳定、可靠,可以作为相关实验研究的理想的体外实验模型。
Aim To establish a model of SNP-induced dorsal root ganglion（DRG） neuron injury based on the method of purifying DRG neuron primary cultures from the neonatal rats.Method DRG were taken out from the intervertebral foramen of neonatal rats and separated into single-celled suspension liquid using collagenase-Dispase,5-FUDR were added to purify the cells,MEM-F12 medium were used to cultivate the cells and NSE immunohistochemical method were applied to identify the DRG neurons.DRG neurons were treated with sodium nitroprusside （SNP） at the eighth day with different concentrations （500,50,5μmol·L-1）.Cells were measured by MTT assay and trypan blue viability test.Results Isolated and cultured DRG neurons survived and grew nicely more than two weeks in the feeding medium,and the purified rate of DRG neurons could be above 90%.The increase of DRG neuron death rate was proportional to the rise of the concentration of the SNP.Conclusion This method is of high stability and efficiency.The neuronal population can be better purified with 5-FUDR.The DRG injury neuron model with SNP preparation could be used as an ideal in vitro model in the relevant experimental studies.
出处
《中国药理学通报》
CAS
CSCD
北大核心
2010年第11期1523-1525,共3页
Chinese Pharmacological Bulletin
基金
安徽省科技攻关重点资助项目(07010300179)
