川芎嗪通过AMPK-NFκB信号通路抑制BV-2小胶质细胞促炎性细胞因子基因表达
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摘要
目的观察川芎嗪(tetramethylpyrazine,TMP)对Aβ25-35诱导的BV-2小鼠小胶质细胞促炎性细胞因子基因表达及对一磷酸腺苷激活的蛋白激酶(adenosine monophosphate-activated protein kinase,AMPK)-核因子κB(nuclear factor kappa B,NFκB)信号通路的影响。方法采用Aβ25-35激活BV-2细胞的炎性反应,测定TMP高、中、低剂量(终浓度分别为100、30、10μmol/L)对Aβ25-35诱导的BV-2细胞白细胞介素-1β(interleukin-1β,IL-1β)、白细胞介素-6(interleukin-6,IL-6)、肿瘤坏死因子-α(tumor necrosis factorα,TNF-α)及诱导型一氧化氮合酶(inducible nitric oxide synthase,iNOS)mRNA表达水平的影响。在AMPK激活剂(AICAR)及抑制剂(化合物C)存在下,检测TMP对AMPK及p65(NFκB亚基)磷酸化的影响。结果 TMP高、中剂量可明显抑制BV-2细胞中IL-1β、IL-6、TNF-α和iNOS基因mRNA表达水平(P<0.01),高剂量TMP可激活BV-2细胞中AMPK(P<0.01),并抑制p65磷酸化(P<0.01)。结论 TMP通过激活BV-2细胞中AMPK活性,抑制NFκB活化,进而抑制促炎性细胞因子基因的表达。
Objective To observe the effects of tetramethylpyrazine(TMP)on the gene expression of proinflammatory cytokines induced by Aβ25-35 and the adenosine monophosphate-activated protein kinase(AMPK)/nuclear factor kappa B(NF-κB)pathway in mouse BV-2 microglial cells.Methods BV-2 cells were treated with Aβ25-35 to induce inflammatory response.The mRNA expression of interleukin-1β(IL-1β),interleukin-6(IL-6),tumor necrosis factor-α(TNF-α),and inducible nitric oxide synthase(iNOS)in BV-2 cells were measured in the presence of high-,middle-,or low-dose TMP(100,30,or 10μmol/L).The effects of TMP on the phosphorylation of AMPK and p65(a subunit of NF-κB)were evaluated in the presence of AMPK activator(AICAR)and inhibitor(Compound C).Results High-and middle-dose TMP significantly inhibited the mRNA expression of IL-1β,IL-6,TNF-α,and iNOS in BV-2cells(P<0.01).High-dose TMP activated AMPK in BV-2cells(P<0.01),but inhibited the phosphorylation of p65(P<0.01).Conclusion TMP can inhibit the mRNA expression of proinflammatory cytokines by activating AMPK and inhibiting the activation of NF-κB in BV-2cells.
Objective To observe the effects of tetramethylpyrazine(TMP)on the gene expression of proinflammatory cytokines induced by Aβ25-35 and the adenosine monophosphate-activated protein kinase(AMPK)/nuclear factor kappa B(NF-κB)pathway in mouse BV-2 microglial cells.Methods BV-2 cells were treated with Aβ25-35 to induce inflammatory response.The mRNA expression of interleukin-1β(IL-1β),interleukin-6(IL-6),tumor necrosis factor-α(TNF-α),and inducible nitric oxide synthase(iNOS)in BV-2 cells were measured in the presence of high-,middle-,or low-dose TMP(100,30,or 10μmol/L).The effects of TMP on the phosphorylation of AMPK and p65(a subunit of NF-κB)were evaluated in the presence of AMPK activator(AICAR)and inhibitor(Compound C).Results High-and middle-dose TMP significantly inhibited the mRNA expression of IL-1β,IL-6,TNF-α,and iNOS in BV-2cells(P<0.01).High-dose TMP activated AMPK in BV-2cells(P<0.01),but inhibited the phosphorylation of p65(P<0.01).Conclusion TMP can inhibit the mRNA expression of proinflammatory cytokines by activating AMPK and inhibiting the activation of NF-κB in BV-2cells.
引文
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[3]Akiyama H,McGeer PL.Specificity of mechanisms for plaque removal after Aβimmunotherapy for Alzheimer disease[J].Nat Med,2004,10(2):117-118.
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[5]王玉,万海同,严伟民,等.川芎嗪与葛根素合用对海马神经元损伤后的影响[J]中国中药杂志,2008,33(4):424-427.
[6]Zhang C,Wang SZ,Zuo PP,et al.Effect of tetramethylpyrazine on learning,memory and cholinergic system in D-galactose-lesioned mice[J].Acta Acad Med Sin,2003,25(5):553-556.
[7]赵琳,魏敏杰,何苗,等.川芎嗪对阿尔采末病模型小鼠学习记忆能力的影响及其机制初探[J].中国药理学通报,2008,24(8):1088-1092.
[8]Zhang Z,Wei T,Hou J,et al.Iron-induced oxidative damage and apoptosis in cerebellar granule cells:attenuation by tetramethylpyrazine and ferulic acid[J].Eur J Pharmacol,2003,467(1):41-47.
[9]刘长安,朱洁,那莎,等.川芎嗪对Aβ25-35诱导体外大鼠海马神经元细胞凋亡的影响[J].天然产物研究与开发,2013,25(5):684-688.
[10]Livak KJ,Schmittgen TD.Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T))method[J].Methods,2001,25(4):402-408.
[11]Fang F,Lue LF,Yan S,et al.RAGE-dependent signaling in microglia contributes to neuroinflammation,Aβaccumulation,and impaired learning/memory in a mouse model of Alzheimer's disease[J].FASEB J,2010,24(4):1043-1055.
[12]McGeer PL,Rogers J,McGeer EG.Inflammation,anti-inflammatory agents and Alzheimer disease:the last 12years[J].J Alzheimers Dis,2006,9(3suppl):271-276.
[13]Britschgi M,Wyss-Coray T.Immune cells may fend off Alzheimer disease[J].Nat Med,2007,13(4):408-409.
[14]Saha RN,Pahan K.Regulation of inducible nitric oxide synthase gene in glial cells[J].Antioxid Redox Signal,2006,8(5-6):929-947.
[15]Zhang J,Dawson VL,Dawson TM,et al.Nitric oxide activation of poly(ADP-ribose)synthetase in neurotoxicity[J].Science,1994,263(5147):687-689.
[16]Sag D,Carling D,Stout RD,et al.Adenosine 5′-monophosphate-activated protein kinase promotes macrophage polarization to an anti-inflammatory functional phenotype[J].J Immunol,2008,181(12):8633-8641.
[17]Wang LM,Wang YJ,Cui M,et al.A dietary polyphenol resveratrol acts to provide neuroprotection in recurrent stroke models by regulating AMPK and SIRT1signaling,thereby reducing energy requirements during ischemia[J].Eur J Neurosci,2013,37(10):1669-1681.
[18]Turnley AM,Stapleton D,Mann RJ,et al.Cellular distribution and developmental expression of AMP-activated protein kinase isoforms in mouse central nervous system[J].J Neurochem,1999,72(4):1707-1716.
[19]Galindo MF,Ikuta I,Zhu X,et al.Mitochondrial biology in Alzheimer's disease pathogenesis[J].J Neurochem,2010,114(4):933-945.
[20]Hotamisligil GS.Endoplasmic reticulum stress and the inflammatory basis of metabolic disease[J].Cell,2010,140(6):900-917.
[21]Greer EL,Oskoui PR,Banko MR,et al.The energy sensor AMP-activated protein kinase directly regulates the mammalian FOXO3transcription factor[J].J Biol chem,2007,282(41):30107-30119.