丙泊酚对新生大鼠脑皮层星形胶质细胞凋亡的影响
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摘要
目的研究丙泊酚对新生大鼠脑皮层星形胶质细胞凋亡的影响及其机制。方法原代分离和培养AST细胞,用GFAP免疫细胞化学鉴定。不同浓度丙泊酚(0、10、30、90μmol·L-1)作用AST细胞8 h。MMT法检测细胞生存率;Annexin V/PI双染检测细胞凋亡;Western blot法测定线粒体cyt-C漏出;分光光度计法检测caspase-3和caspase-9活性。结果分离培养出原代AST,不同浓度丙泊酚(0、10、30、90μmol·L~(-1))浓度依赖性地降低AST细胞生存率,诱导细胞线粒体cyt-C漏出,上调促凋亡蛋白caspase-3和caspase-9,诱发AST凋亡。结论丙泊酚在体外能诱发新生大鼠脑皮层星形胶质细胞凋亡,其机制可能与丙泊酚诱导线粒体cyt-C释放,激活线粒体凋亡通路有关。
Aim To study the effects of propofol on apoptosis of cortical astrocytes isolated from neonatal rats. Methods Pure astrocytes( AST) were obtained from the cultured cerebral cortical cells and identified by the GFAP stain technology in neonatal rats. AST cells were treated with different concentrations of propofol( 0,10,30,90 μmol·L~(-1)) for 8 hours. Cell viability was measured by MTT method,and the apoptosis was detected by Annexin V/PI double staining.Cytochrome C( cyt-C) leakage was detected by Western blot. Caspase-3 and caspase-9 activities were measured by spectrophotometry. Results AST cells were administered with different concentrations of propofol( 0,10,30,90 μmol·L~(-1)) for 8 hours. It decreased the survival rate in a concentration-dependent manner,induced the leakage of cyt-C in mitochondria,up-regulated the expression of pro-apoptotic protein caspase-3 and caspase-9,and induced AST apoptosis.Conclusion Propofol induces the apoptosis of astrocytes in neonatal rat cortex in vitro,which may be related to the activation of mitochondria apoptosis pathway induced by mitochondrial cyt-C release.
Aim To study the effects of propofol on apoptosis of cortical astrocytes isolated from neonatal rats. Methods Pure astrocytes( AST) were obtained from the cultured cerebral cortical cells and identified by the GFAP stain technology in neonatal rats. AST cells were treated with different concentrations of propofol( 0,10,30,90 μmol·L~(-1)) for 8 hours. Cell viability was measured by MTT method,and the apoptosis was detected by Annexin V/PI double staining.Cytochrome C( cyt-C) leakage was detected by Western blot. Caspase-3 and caspase-9 activities were measured by spectrophotometry. Results AST cells were administered with different concentrations of propofol( 0,10,30,90 μmol·L~(-1)) for 8 hours. It decreased the survival rate in a concentration-dependent manner,induced the leakage of cyt-C in mitochondria,up-regulated the expression of pro-apoptotic protein caspase-3 and caspase-9,and induced AST apoptosis.Conclusion Propofol induces the apoptosis of astrocytes in neonatal rat cortex in vitro,which may be related to the activation of mitochondria apoptosis pathway induced by mitochondrial cyt-C release.
引文
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[13]杨慧,关永源.星型胶质细胞在脑缺血性疾病中的神经元保护机制[J].中国药理学通报,2009,25(3):284-6.Yang H,Guan Y Y.Mechanisms of the neuroprotection of astrocyte during cerebral ischemic disease[J].Chin Pharmacol Bull,2009,25(3):284-6.
[14]Yuan X,Zhang L,Wang S,et al.Mitochondrial apoptosis and autophagy in the process of adipose-derived stromal cell differentiation into astrocytes[J].Cell Biol Int,2016,40(2):156-65.
[15]Sun W C,Liang Z D,Pei L.Propofol-induced rno-miR-665 targets BCL2L1 and influences apoptosis in rodent developing hippocampal astrocytes[J].Neurotoxicology,2015,51:87-95.
[2]Germann A L,Shin D J,Manion B D,et al.Activation and modulation of recombinant glycine and GABAAreceptors by 4-halogenated analogues of propofol[J].Br J Pharmacol,2016,173(21):3110-20.
[3]赵施施,张科学,包素红,等.丙泊酚对幼小鼠自主活动和学习记忆行为的影响[J].中国药理学通报,2013,29(2):198-201.Zhao S S,Zhang K X,Bao S H,et al.Effects of propofol on spontaneous motor activity and learning behaviors in juvenile mice[J].Chin Pharmacol Bull,2013,29(2):198-201.
[4]邓小园,陈博,刘红亮,等.TNF-α在丙泊酚诱发的神经元凋亡及认知功能障碍中的作用[J].中国药理学通报,2016,32(7):945-9.Deng X Y,Chen B,Liu H L,et al.Role of TNF-αin propofol-induced neuronal apoptosis and long-term cognitive impairment in neonatal rats[J].Chin Pharmacol Bull,2016,32(7):945-9.
[5]李建立,庞鑫鑫,郭洪霞,等.17β-雌二醇通过PI3K-Akt信号通路抑制丙泊酚诱导皮层神经元凋亡[J].中国病理生理杂志,2016,32(1):58-63.Li J L,Pang X X,Guo H X,et al.17β-estradiol protects cortical neurons from propofol-induced apoptosis by activating PI3K-Akt signaling pathway[J].Chin J Pathophysiol,2016,32(1):58-63.
[6]乔圆,廖雁,南方,等.组胺对星形胶质细胞Egr-1表达的调节作用[J].中国病理生理杂志,2016,32(4):680-5.Qiao Y,Liao Y,Nan F,et al.Effects of histamine on mRNA expression of Egr-1 in astrocytes[J].Chin J Pathophysiol,2016,32(4):680-5.
[7]Hansen D B,Garrido-Comas N,Salter M,et al.HCO3(-)-independent p H regulation in astrocytes in situ is dominated by V-ATPase[J].J Biol Chem,2015,290(13):8039-47.
[8]Chen Z,Wang R,Wu J,et al.Low-dose carbon monoxide inhalation protects neuronal cells from apoptosis after optic nerve crush[J].Biochem Biophys Res Commun,2016,469(4):809-15.
[9]Kim J,Kim S Y,Lee J H,et al.Low-dose dexmedetomidine reduces emergence agitation after desflurane anaesthesia in children undergoing strabismus surgery[J].Yonsei Med J,2014,55(2):508-16.
[10]尚游,吴艳,姚尚龙,等.丙泊酚对大鼠空间学习记忆能力影响的实验研究[J].中国药理学通报,2007,23(6):764-8.Shang Y,Wu Y,Yao S L,et al.Experimental research on the effect of propofol on spatial learning and memory in rats[J].Chin Pharmacol Bull,2007,23(6):764-8.
[11]Chen B,Deng X,Wang B,et al.Etanercept,an inhibitor of TNF-α,prevents propofol-induced neurotoxicity in the developing brain[J].Int J Dev Neurosci,2016,55:91-100.
[12]Reemst K,Noctor S C,Lucassen P J,et al.The indispensable roles of microglia and astrocytes during brain[J].Front Hum Neurosci,2016,10:566.
[13]杨慧,关永源.星型胶质细胞在脑缺血性疾病中的神经元保护机制[J].中国药理学通报,2009,25(3):284-6.Yang H,Guan Y Y.Mechanisms of the neuroprotection of astrocyte during cerebral ischemic disease[J].Chin Pharmacol Bull,2009,25(3):284-6.
[14]Yuan X,Zhang L,Wang S,et al.Mitochondrial apoptosis and autophagy in the process of adipose-derived stromal cell differentiation into astrocytes[J].Cell Biol Int,2016,40(2):156-65.
[15]Sun W C,Liang Z D,Pei L.Propofol-induced rno-miR-665 targets BCL2L1 and influences apoptosis in rodent developing hippocampal astrocytes[J].Neurotoxicology,2015,51:87-95.