摘要
目的研究在星形胶质细胞内加入尼古丁后能否激活细胞内α7尼古丁乙酰胆碱能受体（n AChR）以及激活的α7n AChR对β-淀粉样蛋白（Aβ）的降解作用。方法分离新生乳鼠大脑皮质培养原代星形胶质细胞并纯化、传代,使用细胞免疫荧光法鉴定细胞的纯度。凝胶银染方法鉴定寡聚体聚集情况。试验分为正常对照组、激动剂组、拮抗剂组以及Aβ组激动剂＋Aβ组、拮抗剂＋Aβ组,用蛋白印迹法检测α7n AChR蛋白表达情况以及激活细胞内受体后对Aβ的降解作用。结果经过传代培养细胞得以纯化。体外制备好的Aβ1~42除了含少量的未聚集的单体（4 k D）外,主要以二聚体、三聚体、九聚体等不同聚集形式的寡聚体存在。尼古丁组与对照组相比α7n AChR表达明显升高（P〈0.01）,尼古丁组与拮抗剂组相比α7n AChR表达升高（P〈0.05）;收集的蛋白与上清液,尼古丁组与Aβ组相比Aβ的表达均明显降低（P〈0.01）,激动剂＋Aβ组与拮抗剂＋Aβ组相比Aβ的表达均明显降低（P〈0.01）。结论尼古丁可以激活星形胶质细胞内的α7n AChR,激活的α7n AChR对Aβ具有明显的降解作用,能够为预防和治疗阿尔茨海默氏病（AD）提供一个可能的方向。
Objective To investigate whether α7 neuronal nicotinic acetylcholine receptor（ n AChR） of astrocytes of astrocytes were activated by nicotine,and the degradation effect of α7n AChR on β-amyloid. Methods Primary culture of rat astrocytes were prepared from the cortex and hippocampus of the brains of newborn Sprague-Dawley（ SD） rats. Subculture for three or four times,the passaged cells were demonstrated purity by immunocytochemical stainings with anti-GFAP antibody. Aβ1 ~ 42 oligomer was prepared by chemically synthetic Aβ1 ~ 42 polypeptide in vitro and then the oligomers were identified by sliver staining. The experimental groups were divided into normal control,agonist,blocking agent,or（ and） Aβ groups. The expression of α7n AChR protein and the degradation of Aβ after activation of astrocytic α7 nicotinic receptor were detected by Western blot. Results In addition to discovery of several monomers,low molecular weight and high molecular weight in Aβ oligomers indicated the presence of dimer,trimer and nonamer. Compared with that of control group,the expression of α7n AChR of Nicotine group was significantly higher（ P〈0. 01）,and the expression of α7n AChR of Nicotine group was higher than that of blocking agent group（ P〈0. 05）. Both culture medium and cell lysis,compared with those of Aβ group,the expressions of Aβin Nicotine group of were significantly lower（ P〈0. 01）,and they were significantly lower in Nicotine group than those of blocking agent group（ P〈0. 01）. Conclusions Nicotine could activate the α7n AChR in astrocytes and the activation of α7n AChR has a significant degradation effect on Aβ,which provides a possible direction for the prevention and treatment of AD.
出处
《中国老年学杂志》
CAS
CSCD
北大核心
2016年第13期3112-3115,共4页
Chinese Journal of Gerontology
基金
国家自然科学基金(81260173)
教育部科学技术研究项目(2013032A)
贵州省科技厅国际合作项目(黔科合外G字〔2011〕7014号)
贵州省科技计划(黔科合重大专项字〔2014〕6008号)
