CAJ | 학술논문

背景:在阿尔茨海默病中起着重要作用的淀粉样β蛋白能够诱导细胞凋亡.目的:探讨奥氮平对淀粉样β蛋白25～35诱导的PC12细胞凋亡的机制及其保护作用.单位:解放军总医院南楼神经科.设计:随机设计.材料:实验于2002-05/2003-03在加拿大萨斯卡彻温大学医学院神经精神研究所完成.方法:P12细胞在RPMI1640培养液中培养.在96孔板每孔中接种100μL细胞悬液,在胶原被覆的25 cm2培养瓶中接种5 mL细胞悬液,培养24 h后,分别加50 μmol/L、100 μmol/L奥氮平培养24 h,再加不同浓度的淀粉样β蛋白25～35(0.01 μmol/L、2μmol/L、20μmol/L)培养24h.将收获好的96孔板以淀粉样β蛋白25～35诱导PC12细胞凋亡,采用MTT比色分析测定细胞存活率.收获25 cm2培养瓶中的PC12细胞,应用Westernblot检测奥氮平对PC12细胞Bax、半胱氨酸天冬氨酸蛋白酶3表达的影响.主要观察指标:细胞存活率的测定,PC12细胞中Bax、半胱氨酸天冬氨酸蛋白酶3的表达水平.结果:①细胞存活率比较:淀粉样β蛋白25～35诱导的PC12细胞的细胞活性从75%降低到35%;50μmol/L、100 μmol/L奥氮平预处理组PC12细胞的活性明显提高.②奥氮平对淀粉样β蛋白25～35诱导的PC12细胞凋亡中Bax表达的影响:0.01μmol/L,2 μmol/L,20 μmol/L淀粉样β蛋白25～35处理的PC12细胞Bax的表达增加,50μmol/L奥氮平预处理使淀粉样β蛋白25～35诱导的PC12细胞Bax的表达减低.③奥氮平对Aβ25～35诱导的PC12细胞凋亡中半胱氨酸天冬氨酸蛋白酶3表达的影响:0.001 μmol/L、0.01 μmol/L淀粉样β蛋白处理的PC12无变化,50μmol/L奥氮平预处理对PC12细胞表达亦无影响;2 μmol/L、20μmol/L淀粉样β蛋白25～35处理的PC12细胞表达增高,50μmol/L奥氮平预处理能抑制其表达升高.结论:①淀粉样β蛋白25～35能够诱发与细胞凋亡密切相关的Bax、半胱氨酸天冬氨酸蛋白酶3在培养的PC12细胞中高表达.②奥氮平具有降低其表达,提高PC12细胞存活的保护作用. 揹景:在阿爾茨海默病中起著重要作用的澱粉樣β蛋白能夠誘導細胞凋亡.目的:探討奧氮平對澱粉樣β蛋白25～35誘導的PC12細胞凋亡的機製及其保護作用.單位:解放軍總醫院南樓神經科.設計:隨機設計.材料:實驗于2002-05/2003-03在加拿大薩斯卡徹溫大學醫學院神經精神研究所完成.方法:P12細胞在RPMI1640培養液中培養.在96孔闆每孔中接種100μL細胞懸液,在膠原被覆的25 cm2培養瓶中接種5 mL細胞懸液,培養24 h後,分彆加50 μmol/L、100 μmol/L奧氮平培養24 h,再加不同濃度的澱粉樣β蛋白25～35(0.01 μmol/L、2μmol/L、20μmol/L)培養24h.將收穫好的96孔闆以澱粉樣β蛋白25～35誘導PC12細胞凋亡,採用MTT比色分析測定細胞存活率.收穫25 cm2培養瓶中的PC12細胞,應用Westernblot檢測奧氮平對PC12細胞Bax、半胱氨痠天鼕氨痠蛋白酶3錶達的影響.主要觀察指標:細胞存活率的測定,PC12細胞中Bax、半胱氨痠天鼕氨痠蛋白酶3的錶達水平.結果:①細胞存活率比較:澱粉樣β蛋白25～35誘導的PC12細胞的細胞活性從75%降低到35%;50μmol/L、100 μmol/L奧氮平預處理組PC12細胞的活性明顯提高.②奧氮平對澱粉樣β蛋白25～35誘導的PC12細胞凋亡中Bax錶達的影響:0.01μmol/L,2 μmol/L,20 μmol/L澱粉樣β蛋白25～35處理的PC12細胞Bax的錶達增加,50μmol/L奧氮平預處理使澱粉樣β蛋白25～35誘導的PC12細胞Bax的錶達減低.③奧氮平對Aβ25～35誘導的PC12細胞凋亡中半胱氨痠天鼕氨痠蛋白酶3錶達的影響:0.001 μmol/L、0.01 μmol/L澱粉樣β蛋白處理的PC12無變化,50μmol/L奧氮平預處理對PC12細胞錶達亦無影響;2 μmol/L、20μmol/L澱粉樣β蛋白25～35處理的PC12細胞錶達增高,50μmol/L奧氮平預處理能抑製其錶達升高.結論:①澱粉樣β蛋白25～35能夠誘髮與細胞凋亡密切相關的Bax、半胱氨痠天鼕氨痠蛋白酶3在培養的PC12細胞中高錶達.②奧氮平具有降低其錶達,提高PC12細胞存活的保護作用.
배경:재아이자해묵병중기착중요작용적정분양β단백능구유도세포조망.목적:탐토오담평대정분양β단백25～35유도적PC12세포조망적궤제급기보호작용.단위:해방군총의원남루신경과.설계:수궤설계.재료:실험우2002-05/2003-03재가나대살사잡철온대학의학원신경정신연구소완성.방법:P12세포재RPMI1640배양액중배양.재96공판매공중접충100μL세포현액,재효원피복적25 cm2배양병중접충5 mL세포현액,배양24 h후,분별가50 μmol/L、100 μmol/L오담평배양24 h,재가불동농도적정분양β단백25～35(0.01 μmol/L、2μmol/L、20μmol/L)배양24h.장수획호적96공판이정분양β단백25～35유도PC12세포조망,채용MTT비색분석측정세포존활솔.수획25 cm2배양병중적PC12세포,응용Westernblot검측오담평대PC12세포Bax、반광안산천동안산단백매3표체적영향.주요관찰지표:세포존활솔적측정,PC12세포중Bax、반광안산천동안산단백매3적표체수평.결과:①세포존활솔비교:정분양β단백25～35유도적PC12세포적세포활성종75%강저도35%;50μmol/L、100 μmol/L오담평예처리조PC12세포적활성명현제고.②오담평대정분양β단백25～35유도적PC12세포조망중Bax표체적영향:0.01μmol/L,2 μmol/L,20 μmol/L정분양β단백25～35처리적PC12세포Bax적표체증가,50μmol/L오담평예처리사정분양β단백25～35유도적PC12세포Bax적표체감저.③오담평대Aβ25～35유도적PC12세포조망중반광안산천동안산단백매3표체적영향:0.001 μmol/L、0.01 μmol/L정분양β단백처리적PC12무변화,50μmol/L오담평예처리대PC12세포표체역무영향;2 μmol/L、20μmol/L정분양β단백25～35처리적PC12세포표체증고,50μmol/L오담평예처리능억제기표체승고.결론:①정분양β단백25～35능구유발여세포조망밀절상관적Bax、반광안산천동안산단백매3재배양적PC12세포중고표체.②오담평구유강저기표체,제고PC12세포존활적보호작용.
BACKGROUND: β-amyloid has been proved to be capable of inducing cell apoptosis and play a vital role in Alzheimer disease (AD).OBJECTIVE: To probe into olanzapine's protective effect and mechanism of PC12 cell apoptosis induced by β-amyloid 25-35.SETTING: Department of Neurology, Southern Building of the General Hospital of Chinese PLA.DESIGN: Randomized design.MATERIALS: This experiment was carried out in the Neuropsychopathic Research Institute, Medical College of the University of Saskatchewan(Canada), between May 2002 and March 2003.METHODS: P12 cells were cultured with RPMI1640 culture medium.100 μL cell suspension was inoculated in each well of 96-well culture plate, and 5 mL suspension was inoculated in 25 cm2 culture bottle covered with collagen and cultured for 24 hours, then with additional 50 μmol/L and 100 μmol/L olanzapine, respectively, for 24 hours, and β-amyloid 25-35 of different concentrations (0.01 μmol/L, 2 μmol/L and 20 μmol/L) for 24 hours. PC12 cell apoptosis was induced by β-amyloid 25-35 in 96-well culture plate and cells were harvested to assay their survival rate with MTF colorimetric assay. PC12 cells in 25-cm2 culture bottles were also harvested to detect the effect of olanzapine on Bax and caspase-3 expression in PC12 cells using Western blot assay.MAIN OUTCOME MEASURES: ① Cell survival rate; ② the expression of Bax and caspase-3 in PC12 cells.RESULTS: ① Cell survival rate: cell activity was found declined from 75% to 35% in PC12 cells induced by β-amyloid 25-35, but obviously increased in PC12 cells due to pretreatment with olanzapine of 50 μmol/L and 100 μmol/L. ② Olanzapine's effects on Bax expression in PC12 cell apoptosis induced by β-amyloid 25-35: Bax expression increased in PC12cells due to exposure to β-amyloid 25-35 of 0.01 μmol/L, 2 μmol/L and 20 μmol/L, but it could be suppressed if pretreated with olanzapine of 50 μmol/L. ③ Effect of olanzapine on caspase-3 expression in PC12apoptotic cells induced by β-amyloid 25-35: There was no change in PC12cells induced by 0.001 μmol/L or 0.01 μmol/L of β-amyloid, as well as in PC12 cells pretreated with 50 μmol/L olanzapine. However, caspase-3 expression obviously increased in PC12 cells exposed to 2 μmol/L and 20 μmol/L of β-amyloid 25-35, and it could be suppressed by pretreatment with 50 μmol/L of olanzapine.CONCLUSION: ① β-amyloid 25-35 can induce the high expression of cell apoptosis related Bax and caspase-3 in vitro cultured PC12 cells. ②Olanzapine can reduce the expression, thus enhancing the survival rate of PC 12 cells.