【摘要】 目的研究蛋白酶体抑制剂PSI诱导PC12细胞胞浆内嗜酸性包涵体的形成及演变过程。方法取进入对数生长期的PC12细胞。孵育24h后,加入PSI(终浓度10μmol/L),分别于给药后3、6、9、12、24、48、72、96、120h取出细胞并甩片固定,行HE染色及α-synuclein免疫组织化学染色,光镜下观察形态变化。结果HE染色:PSI作用6h后,PC12细胞胞浆呈现嗜伊红染色,胞体变大;至12h可见明显的嗜伊红聚集的核心;24h后,几乎所有的细胞都形成了核旁嗜伊红球状包涵体;其后,胞核及胞浆逐渐缺失,于120h仅余聚集的嗜伊红球状包涵体。免疫组化染色:PSI作用3h后,可见到散在于整个胞浆的α-synuclein免疫反应阳性颗粒;6h后,细胞明显增大,胞浆中颗粒增多;12h后,颗粒聚集更加明显,形成明显的聚集中心;24h后,几乎所有的细胞都形成了α-synuclein免疫反应阳性的球状包涵体;随时间延长,胞核及胞浆逐渐缺失,至120h仅见固缩的α-synuclein免疫反应阳性球状包涵体。结论PSI作用于PC12细胞,早期出现散在于胞浆中的嗜酸性和α-synuclein免疫反应阳性的点状颗粒,然后这些颗粒进一步在核旁聚集,形成聚集中心,最后形成固缩的包涵体,细胞外包涵体可独立存在。这种包涵体形成和演变过程与以人类PD为代表的神经变性疾病中的包涵体非常相似,可以作为研究包涵体相关问题的有力工具。更多还原

【Abstract】 Objective To study the formation and development of inclusions in dopaminergic PC12 cells treated with proteasome inhibitor PSI.Methods PC12 cells were incubated for 24 hours,then were treated with 10μmol/L PSI for 3,6,9,12,24,48,72,96,120hours respectively,fixed with formaldehyde,stained with H&E method andα-synuclein immunohistochemistry.Results At 3h,α-synuclein immunoreactive granules were found throughout the cytoplasm;at 6h,the number ofα-synuclein immunoreactive granules increased;at 12h,eosinophilic materials andα-synuclein immunoreactive granules concentrated and formed inclusion closed to nucleus;at 24h,in- clusions were seen in almost all PC12 cells;at 120h,the nucleus and cytoplasm decomposed,only inclusions present- ed.Conclusion Treated PC12 cells with proteasome inhibitor can produce cytoplasmic eosinophilic andα-synucle- in immunoreactive inclusions.The formation and development of inclusions in this model are consistent with those in human Parkinson disease and defused Lewy body disease.This model is helpful to further explore the mecha- nism of neurodegenerative diseases.更多还原