摘要
目的探究自噬激活在丙泊酚诱导海马神经元凋亡中的作用。方法取胎鼠提取分离原代海马神经元,随机分为对照组、丙泊酚组、雷帕霉素组与氯喹组。采用二甲氧唑黄比色法(XTT)和流式细胞术检测神经元凋亡,Western blot法检测自噬与凋亡相关蛋白表达。取老龄大鼠随机分为对照组、丙泊酚组、雷帕霉素组与氯喹组。以100 mg/kg丙泊酚连续麻醉1周,期间以50 mg/kg雷帕霉素和10 mg/kg氯喹治疗。采用Morris水迷宫检测大鼠认知功能,TUNEL染色检测海马神经元凋亡情况,高尔基染色观察海马神经元自噬情况,Western blot法检测自噬与凋亡相关蛋白表达。结果离体试验中,雷帕霉素逆转了丙泊酚造成的细胞凋亡。与丙泊酚组相比,雷帕霉素组微管相关蛋白轻链3Ⅱ/微管相关蛋白轻链3Ⅰ(LC3Ⅱ/LC3Ⅰ)与Beclin-1表达增加,但凋亡蛋白剪切的半胱氨酸蛋白酶-3(Cleaved-caspase-3)与Bax表达降低。在体研究中,与丙泊酚组相比,雷帕霉素组水迷宫逃避潜伏期减少,且雷帕霉素组TUNEL阳性细胞数与自噬小体数量减少。此外,雷帕霉素组哺乳动物雷帕霉素靶蛋白(mTOR)表达降低,且LC3Ⅱ/LC3Ⅰ与Beclin-1表达增加,Cleaved-caspase-3与Bax表达降低。但氯喹对自噬和凋亡相关蛋白没有影响。结论雷帕霉素通过抑制mTOR信号活化,进一步激活了细胞自噬,改善了丙泊酚造成的神经元凋亡,提高了大鼠认知功能。
Objective To explore the effects of autophagy activation on propofol-induced apoptosis in hippocampal neurons.Methods Primary hippocampal neurons were extracted and isolated from fetal rats and randomly divided into control group,propofol group,rapamycin group and chloroquine group.2,3-Bis-(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide(XTT)and flow cytometry were used to detect apoptosis,and Western blot was used to detect the expression of autophagy and apoptosis-related proteins.Then,aged rats were randomly divided into control group,propofol group,rapamycin group and chloroquine group.100 mg/kg propofol was used foRcontinuous anesthesia foR1 week,during which 50 mg/kg rapamycin and 10 mg/kg chloroquine were treated.Morris wateRmaze was used to detect cognitive function,TUNEL staining was used to detect apoptosis,Golgi staining was used to observe autophagy,and Western blot was used to detect autophagy-related apoptosis-related protein expression.Results In vitro,rapamycin obviously reversed the apoptosis caused by propofol,but chloroquine had no effect.Compared with propofol group,the expression of microtubule-associated protein light chain 3Ⅱ/microtubule-associated protein light chain 3 I(LC3Ⅱ/LC3Ⅰ)and Beclin-1 in the rapamycin group increased,but the expression of apoptotic proteins Cleaved-caspase-3 and Bax decreased.In vivo,compared with propofol group,the wateRmaze escape latency of the rapamycin group reduced.In addition,the numbeRof TUNEL-positive cells and autophagosomes in the rapamycin group decreased.Furthermore,the expression of mammalian target of rapamycin(mTOR)in the rapamycin group decreased,and the expression of LC3Ⅱ/LC3Ⅰand Beclin-1 increased,as well as the expression of Cleaved-caspase-3 and Bax decreased.But chloroquine had no effect on autophagy and apoptosis-related proteins.Conclusion Rapamycin can furtheRactivate autophagy by inhibiting the activation of mTORsignal,ameliorate the neuronal apoptosis caused by propofol,which will lead to the improvement of the
作者
梅静
喇宏玲
徐桂萍
Mei Jing;La Hongling;Xu Guiping(Dept of Anesthesiology,People′s Hospital of Xinjiang UyguRAutonomous Region,Urumqi 830001)
出处
《安徽医科大学学报》
CAS
北大核心
2022年第10期1552-1558,共7页
Acta Universitatis Medicinalis Anhui
基金
新疆维吾尔自治区自然科学基金(编号:2019D01C116)。
关键词
丙泊酚
自噬
神经凋亡
认知功能
雷帕霉素
propofol
autophagy
neuroapoptosis
cognitive function
rapamycin
