阿魏酸对H_2O_2致PC12细胞氧化损伤的保护作用
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摘要
目的:探讨阿魏酸(ferulic acid,FA)对H_2O_2诱导PC12细胞氧化损伤的保护作用及其作用机制。方法:H_2O_2诱导PC12细胞构建氧化应激损伤模型,不同剂量阿魏酸(终浓度为1,10,100μmol·L~(-1))进行干预;采用噻唑蓝(MTT)比色法检测细胞存活率,生化法检测细胞上清液中乳酸脱氢酶(LDH)和丙二醛(MDA)的含量以及细胞中超氧化物歧化酶(SOD)的活性,采用实时荧光定量聚合酶链式反应(Real-time PCR)检测细胞中胰岛素样生长因子-1(IGF-1) mRNA的表达,蛋白免疫印迹法(Western blot)检测IGF-1蛋白的表达。结果:不同剂量的阿魏酸(终浓度为1,10,100μmol·L~(-1))干预24 h后能明显改善H_2O_2诱导的PC12细胞的氧化损伤,与模型组比较,给药组细胞存活率明显升高,细胞上清液中LDH的漏出率明显减少(P <0. 01); MDA的含量明显降低(P <0. 05),同时细胞中SOD的含量显著升高(P <0. 01); Real-time PCR结果显示,与空白组比较,模型组IGF-1 mRNA显著降低(P <0. 01),与模型组比较,药物干预24 h后IGF-1 mRNA表达显著升高(P <0. 01); Western blot结果显示,与空白组比较,模型组IGF-1蛋白的表达显著降低(P <0. 01),与模型组比较,药物作用24 h后,能上调IGF-1蛋白的表达(P <0. 01)。结论:FA能够抑制H_2O_2诱导的PC12细胞的氧化损伤,其保护作用机制可能与胰岛信号通路相关。
Objective: To investigate the protective effect of ferulic acid on PC12 cells injured by H_2O_2 and the molecular mechanisms. Method: The oxidative stress model was established by treating PC12 cells with H_2O_2,and then different dosages of ferulic acid( 1,10,100 μmol·L~(-1)) were used for intervention. Methyl thiazolyl tetrazolium( MTT) assay was used to evaluate the cell viability, lactate dehydrogenase( LDH) and malondialdelyde( MDA) in cell supernatant,and superoxidedismutase( SOD) in cells was tested by biochemical method respectively. Insulin-like growth factors-1( IGF-1) mRNA and protein expressions were detected by Realtime fluorescence quantitative polymerase chain reaction( Real-time PCR) and Western blot. Result: The 24 h intervention with different dosages of ferulic acid( 1,10,100 μmol·L~(-1)) could significantly improve the oxidative damage of PC12 cells induced by H_2O_2,compared with the model group,ferulic acid at 1,10,100 μmol·L~(-1) significantly increased PC12 cells viability, significantly decreased LDH and MDA content in cell supernatant( P < 0. 05,P < 0. 01),and enhanced SOD activity( P < 0. 01). Real-time PCR and Western blot showed that compared with control group,IGF-1 mRNA and protein expressions in model group decreased significantly( P <0. 01),compared with model group,IGF-1 mRNA and protein expressions in ferulic acid group increased significantly( P < 0. 01). Conclusion: Ferulic acid exerts a protective effect on H_2O_2-inducing PC12 cells injury,which might be related to insulin signaling pathways.
Objective: To investigate the protective effect of ferulic acid on PC12 cells injured by H_2O_2 and the molecular mechanisms. Method: The oxidative stress model was established by treating PC12 cells with H_2O_2,and then different dosages of ferulic acid( 1,10,100 μmol·L~(-1)) were used for intervention. Methyl thiazolyl tetrazolium( MTT) assay was used to evaluate the cell viability, lactate dehydrogenase( LDH) and malondialdelyde( MDA) in cell supernatant,and superoxidedismutase( SOD) in cells was tested by biochemical method respectively. Insulin-like growth factors-1( IGF-1) mRNA and protein expressions were detected by Realtime fluorescence quantitative polymerase chain reaction( Real-time PCR) and Western blot. Result: The 24 h intervention with different dosages of ferulic acid( 1,10,100 μmol·L~(-1)) could significantly improve the oxidative damage of PC12 cells induced by H_2O_2,compared with the model group,ferulic acid at 1,10,100 μmol·L~(-1) significantly increased PC12 cells viability, significantly decreased LDH and MDA content in cell supernatant( P < 0. 05,P < 0. 01),and enhanced SOD activity( P < 0. 01). Real-time PCR and Western blot showed that compared with control group,IGF-1 mRNA and protein expressions in model group decreased significantly( P <0. 01),compared with model group,IGF-1 mRNA and protein expressions in ferulic acid group increased significantly( P < 0. 01). Conclusion: Ferulic acid exerts a protective effect on H_2O_2-inducing PC12 cells injury,which might be related to insulin signaling pathways.
引文
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[19]白宇,贺云,杨丽霞.活血化瘀类中药治疗糖尿病肾病机制的研究进展[J].中国实验方剂学杂志,2018,24(23):200-206.
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[21] Prabhu B M,Medina-Bolivar F,Condori J,et al.Neuroprotective effect of peanut hairy root extract against oxidative stress in PC 12 derived neurons[J]. J Med Active Plants,2013,1:125-133.
[22] Freude S,Schilbach K,Schubert M. The role of IGF-1receptor signaling for the pathogenesis of Alzheimer's disease:from model organisms to human disease[J].Curr Alzheimer Res,2009,6(3):213-223.
[23] Chesik D,de Keyser J,Wilczak N. Insulin-like growth factor system regulates oligodendrolial cell behavior:therapeutic potential in CNS[J]. J Mol Neurosci,2008,35(1):81-90.
[2] LIU Q,HAN L,DU Q,et al. The association between oxidative stress,activator protein-1,inflammatory,total antioxidant status and artery stiffness and the efficacy of olmesartan in elderly patients withmild-to-moderate es sential hypertension[J]. Clin Exp Hypertens,2016,38(4):365-369.
[3] Walker A M,Stevens J J,Ndebele K. et al. Evaluation of arsenic trioxide potential for lung cancer treatment:assessment of apoptotic mechanisms and oxidative damage[J]. Cancer Sci Ther,2016,8(1):1-9.
[4] Sharma A,Bist R, Bubber P. Thiamine deficiency induces oxidative stress in brain mitochondria of Mus musculus[J]. J Physiol Biochem,2013,69(3):539-546.
[5] Behl C. Alzheimer's disease and oxidative stress:implications for novel therapeutic approaches[J]. Prog Neurobiol,1999,57(3):301-323.
[6] Kuang X,CHEN Y S,WANG L F,et al. Klotho upregulation contributes to the neuroprotection of ligustilide in an Alzheimer's disease mouse model[J].Neurobiol Aging,2014,35(1):169-178.
[7] Kawabata K,Yamamoto T,Hara A,et al. Modifying effects offerulic acid on azoxymethane-induced colon carcinogenesis in F344 rats[J]. Cancer Lett,2000,157(1):15-21.
[8]李福龙,李继红,刘艳凯,等.川芎嗪、当归注射液对DIC大鼠血小板功能和器官血流量的影响[J].基础医学与临床,2006,26(8):909-910.
[9]龚婉,陈晓玲,周莉,等.阿魏酸钠对大鼠脑缺血再灌注炎症损伤的保护作用及机制分析[J].中国实验方剂学杂志,2019,25(3):94-99.
[10]黄浩,马增春,王宇光,等.阿魏酸对脂多糖损伤的PC12细胞和大鼠海马神经元细胞的保护作用[J].中国药理学与毒理学杂志,2016,30(4):330-337.
[11] ZHANG B,LI W,DONG M. Flavonoids of kudzu root fermented by eurtotium cristatum protected rat pheochromocytoma line 12(PC12)cells against h2o2-induced apoptosis[J]. Int J Mol Sci, 2017,18(12):e2754.
[12] MA S,LIU X,XUN Q,et al. Neuroprotective effect of ginkgolide k against h2o2-induced PC12 cell cytotoxicity by ameliorating mitochondrial dysfunction and oxidative stress.[J]. Biological,2014,37(2):217-225.
[13] Pohanka M. Role of oxidative stress in infectious diseases[J]. Folia Microbiol,2013,58(6):503-513.
[14] Emerit J, Edeas M, Bricaire F. Neurodegenerative diseases and oxidative stress[J]. Biomed Pharmacother,2004,58(1):39-46.
[15] LIN M T,Beal M F,Mitochondrial dysfunction and oxidative stress in neurodegenerative diseases[J].Nature,2006,443(7113):787-795.
[16] ZHOU T,XU B N,QUE H P,et al. Neurons derived from PC 12 cells have the potential to develop synapse with primary neurons from rat cortex[J]. Acta Neurobiol Exp,2006,66(2):105-112.
[17] CHEN B,YUE R,YANG Y,et al. Protective effects of(E)-2-(1-hydroxyl-4-oxocyclohexyl)ethyl caffeine against hydrogen peroxide-induced injury in PC12 cells[J]. Neurochem Res,2015,40(3):531-541.
[18] HU W,WANG G,LI P,et al. Neuroprotective effects of macranthoin G from Eucommia ulmoides against hydrogen peroxide-induced apoptosis in PC12 cells via inhibiting NF-kappaB activation[J]. Chem Biol Interact,2014,224:108-116.
[19]白宇,贺云,杨丽霞.活血化瘀类中药治疗糖尿病肾病机制的研究进展[J].中国实验方剂学杂志,2018,24(23):200-206.
[20] HONG H, LIU G Q. Protection against hydrogen peroxideinduced cytotoxicity in PC 12 cells by scutellarin[J]. Life Sci,2004,74(24):2959-2973.
[21] Prabhu B M,Medina-Bolivar F,Condori J,et al.Neuroprotective effect of peanut hairy root extract against oxidative stress in PC 12 derived neurons[J]. J Med Active Plants,2013,1:125-133.
[22] Freude S,Schilbach K,Schubert M. The role of IGF-1receptor signaling for the pathogenesis of Alzheimer's disease:from model organisms to human disease[J].Curr Alzheimer Res,2009,6(3):213-223.
[23] Chesik D,de Keyser J,Wilczak N. Insulin-like growth factor system regulates oligodendrolial cell behavior:therapeutic potential in CNS[J]. J Mol Neurosci,2008,35(1):81-90.
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