头穴针刺对局灶性脑缺血大鼠海马旁回炎性反应相关细胞因子表达及脑梗死体积的影响
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摘要
目的:观察头穴针刺干预对局灶性脑缺血大鼠海马旁回白细胞介素(IL)-10、IL-6、IL-1β表达及脑梗死体积的影响,探讨头穴针刺调节急性缺血性脑血管病炎性反应的机制。方法:SD大鼠中随机抽取12只作为正常组,36只采用线栓法建立局灶性脑缺血模型,随机分为模型组、药物组及头针组,每组12只。药物组以吡咯烷二硫代氨基甲酸盐(100mg·kg-1·d-1)腹腔注射,1次/d,共7d;头针组给予双侧"顶颞前斜线"快速透刺治疗,留针20min,1次/d,共7d。分别干预刺前和末次干预结束即刻对各组大鼠进行神经功能缺损评分(NDS)和神经行为学评分(NS)。以2,3,5-三苯基氯化四氮唑(TTC)染色法测定各组大鼠脑梗死体积,免疫组织化学法检测各组大鼠海马旁回IL-10、IL-6、IL-1β的表达。结果:与同组干预前比较,药物组与头针组大鼠NDS和NS均显著降低(P<0.01)。与正常组比较,模型组大鼠NDS和NS升高(P<0.01),脑梗死体积明显增大(P<0.01),海马旁回IL-10、IL-6、IL-1β的表达均明显增高(P<0.01);与模型组比较,药物组和头针组大鼠NDS、NS及海马旁回IL-6、IL-1β表达均明显降低(P<0.05),脑梗死体积明显缩小(P<0.05),头针组IL-10表达明显增高(P<0.05)。结论:上调IL-10的表达,抑制IL-6、IL-1β等促炎细胞因子的过度表达,可能是头穴针刺调节缺血性脑血管病炎性反应的机制之一。
Objective To observe the influence of scalp acupuncture on cerebral infarct size and expression of IL-10,IL-6,and IL-1βin the para-hippocampal gyrus in acute ischemic cerebrovascular disease(AICD)rats,so as to investigate its mechanisms underlying improvement of AICD.Methods Forty-eight male SD rats were randomly allocated to normal control(control),AICD model,medication,and scalp acupuncture groups(n=12 per group).The AICD model was established by occlusion of the middle cerebral artery(MCAO).Rats of the medication group received intraperitoneal injection of Ammonium1-Pyrrolidinedithiocarbamate(APDC,100 mg·kg-1·d-1),once daily for 7 days.Scalp acupuncture stimulation was applied to bilateral"Dingnieqianxiexian"(MS6)once daily for 7 days.Before and after intervention,the neurologic deficit score(NDS)and the neurological score(NS)were evaluated according to Longa's and Schbitz's methods,respectively.At the end of the intervention,the para-hippocampal gyrus and whole brain were collected respectively.The expression levels of IL-10,IL-6 and IL-1βin the para-hippocampal gyrus tissue were detected by immunohistochemistry,and the cerebral infarct volume of the brain was detected by triphenyltetrazollium chloride(TTC)staining after sectioning.Results Following modeling,the NDS,NS and the expression of IL-10,IL-6 and IL-1βin para-hippocampal gyrus were significantly increased in the model group compared with the control group(P<0.01).After the intervention,the NDS,NS and infarct volume,and the expression levels of IL-6 and IL-1βin the para-hippocampal gyrus were significantly decreased in both medication and scalp acupuncture groups compared with the model group(P<0.05),and the expression of IL-10 was further obviously up-regulated in the scalp acupuncture group(P<0.05)rather than in the medication group(P>0.05).The effect of scalp acupuncture was obviously superior to that of medication in up-regulating IL-10 expression level(P<0.05).No significant differences were found between the medication and scalp acupuncture groups in the levels of NDS,NS,infarct volume,IL-6 and IL-1βproteins(P>0.05).Conclusion Scalp acupuncture can improve neurological function and reduce infarct volume in AICD rats,which may be associated with its function in up-regulating the expression of IL-10 and in inhibiting the expression of IL-6 and IL-1βto reduce inflammation reaction.
Objective To observe the influence of scalp acupuncture on cerebral infarct size and expression of IL-10,IL-6,and IL-1βin the para-hippocampal gyrus in acute ischemic cerebrovascular disease(AICD)rats,so as to investigate its mechanisms underlying improvement of AICD.Methods Forty-eight male SD rats were randomly allocated to normal control(control),AICD model,medication,and scalp acupuncture groups(n=12 per group).The AICD model was established by occlusion of the middle cerebral artery(MCAO).Rats of the medication group received intraperitoneal injection of Ammonium1-Pyrrolidinedithiocarbamate(APDC,100 mg·kg-1·d-1),once daily for 7 days.Scalp acupuncture stimulation was applied to bilateral"Dingnieqianxiexian"(MS6)once daily for 7 days.Before and after intervention,the neurologic deficit score(NDS)and the neurological score(NS)were evaluated according to Longa's and Schbitz's methods,respectively.At the end of the intervention,the para-hippocampal gyrus and whole brain were collected respectively.The expression levels of IL-10,IL-6 and IL-1βin the para-hippocampal gyrus tissue were detected by immunohistochemistry,and the cerebral infarct volume of the brain was detected by triphenyltetrazollium chloride(TTC)staining after sectioning.Results Following modeling,the NDS,NS and the expression of IL-10,IL-6 and IL-1βin para-hippocampal gyrus were significantly increased in the model group compared with the control group(P<0.01).After the intervention,the NDS,NS and infarct volume,and the expression levels of IL-6 and IL-1βin the para-hippocampal gyrus were significantly decreased in both medication and scalp acupuncture groups compared with the model group(P<0.05),and the expression of IL-10 was further obviously up-regulated in the scalp acupuncture group(P<0.05)rather than in the medication group(P>0.05).The effect of scalp acupuncture was obviously superior to that of medication in up-regulating IL-10 expression level(P<0.05).No significant differences were found between the medication and scalp acupuncture groups in the levels of NDS,NS,infarct volume,IL-6 and IL-1βproteins(P>0.05).Conclusion Scalp acupuncture can improve neurological function and reduce infarct volume in AICD rats,which may be associated with its function in up-regulating the expression of IL-10 and in inhibiting the expression of IL-6 and IL-1βto reduce inflammation reaction.
引文
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[14]SCHBITZ W R,BERGER C,KOLLMAR R,et al.Effect of brain-derived neurotrophic factor treatment and forced arm use on functional motor recovery after small cortical ischemia[J].Stroke,2004,35(4):992-997.
[15]ZGAVC T,CEULEMANS A G,HACHIMI-IDRISSI S,et al.The neuroprotective effect of post ischemic brief mild hypothermic treatment correlates with apoptosis,but not with gliosis in endothelin-1treated rats[J].BMC Neurosci,2012,13:105.
[16]CEULEMANS A G,ZGAVC T,KOOIJMAN R,et al.Mild hypothermia causes differential,time-dependent changes in cytokine expression and gliosis following endothelin-1-induced transient focal cerebral ischemia[J].J Neuroinflammation,2011,8:60.
[17]DI NAPOLI M,SHAH I M.Neuroinflammation and cerebrovascular disease in old age:a translational medicine perspective[J].J Aging Res,2011,2011:857484.
[18]LIU F D,MCCULLOUGH L D.Inflammatory responses in hypoxic ischemic encephalopathy[J].Acta Pharmacol Sin,2013,34(9):1121-1130.
[19]ORION D,SCHWAMMENTHAL Y,RESHEF T,et al.Interleukin-6and soluble intercellular adhesion molecule-1in acute brain ischaemia[J].Eur J Neurol,2008,15(4):323-328.
[20]PRADILLO J M,DENES A,GREENHALGH A D,et al.Delayed administration of interleukin-1receptor antagonist reduces ischemic brain damage and inflammation in comorbid rats[J].J Cereb Blood Flow Metab,2012,32(9):1810-1819.
[21]CASO J R,MORO M A,LORENZO P,et al.Involvement of IL-1βin acute stress-induced worsening of cerebral ischaemia in rats[J].Eur Neuropsychopharmacol,2007,17(9):600-607.
[22]SRTLE K,ZHOU J H,SHEN W H,et al.Interleukin-10in the brain[J].Crit Rev Immunol,2001,21(5):427-449.
[23]GRILLI M,BARBIERI I,BASUDEV H,et al.Interleukin-10modulates neuronal threshold of vulnerability to ischaemic damage[J].Eur J Neurosci,2000,12(7):2265-2272.
[24]GARCIA J M,STILLINGS S A,LECLERC J L,et al.Role of interleukin-10in acute brain injuries[J].Front Neurol,2017,8:244.
[25]VAN EXEL E,GUSSEKLOO J,DE CRAEN A J,et al.Inflammation and stroke:the Leiden 85-plus study[J].Stroke,2002,33(4):1135-1138.
[26]PREZ-DE PUIG I,MIRF,SALAS-PERDOMO A,et al.IL-10deficiency exacerbates the brain inflammatory response to permanent ischemia without preventing resolution of the lesion[J].J Cereb Blood Flow Metab,2013,33(12):1955-1966.
[27]HUXFORD T,GHOSH G.A structural guide to proteins of the NF-κB signaling module[J].Cold Spring Harb Perspect Biol,2009,1(3):a000075.
[28]DRIESSLER F,VENSTROM K,SABAT R,et al.Molecular mechanisms of interleukin-10-mediated inhibition of NF-κB activity:a role for p50[J].Clin Exp Immunol,2004,135(1):64-73.
[29]PFEILSCHIFTER W,CZECH B,HOFFMANN B P,et al.Pyrrolidine dithiocarbamate activates p38MAPK and protects brain endothelial cells from apoptosis:a mechanism for the protective effect in stroke[J].Neurochem Res,2010,35(9):1391-1401.
[30]WANG Z X,SHAN W R,CAO J B,et al.Early administration of pyrrolidine dithiocarbamate extends the therapeutic time window of tissue plasminogen activator in a male rat model of embolic stroke[J].J Neurosci Res,2018,96(3):449-458.
[2]LIU J,CHEN Q X,JIAN Z H,et al.Daphnetin protects against cerebral ischemia/reperfusion injury in mice via inhibition of TLR4/NF-κB signaling pathway[J].Biomed Res Int,2016,2016:2816056.
[3]MONTANER J,ROVIRA A,MOLINA C A,et al.Plasmatic level of neuroinflammatory markers predict the extent of diffusion-weighted image lesions in hyperacute stroke[J].J Cereb Blood Flow Metab,2003,23(12):1403-1407.
[4]WONG A A,DAVIS J P,SCHLUTER P J,et al.The time course and determinants of temperature within the first 48hafter ischaemic stroke[J].Cerebrovasc Dis,2007,24(1):104-110.
[5]AMARO S,CHAMORRO.Translational stroke research of the combination of thrombolysis and antioxidant therapy[J].Stroke,2011,42(5):1495-1499.
[6]BACHIS A,COLANGELO A M,VICINI S,et al.Interleukin-10prevents glutamate-mediated cerebellar granule cell death by blocking caspase-3-like activity[J].J Neurosci,2001,21(9):3104-3112.
[7]王金海,张星华,杜小正,等.头针治疗缺血性卒中疗效的Meta分析[J].中国循证医学杂志,2015,15(10):1161-1167.
[8]LIU H,SUN X W,ZOU W,et al.Scalp acupuncture attenuates neurological deficits in a rat model of hemorrhagic stroke[J].Complement Ther Med,2017,32:85-90.
[9]王金海,赵敏,鲍英存,等.头穴透刺对急性脑梗死患者血清超敏C反应蛋白及炎性反应因子水平的影响[J].针刺研究,2016,41(1):80-84.
[10]LONGA E Z,WEINSTEIN P R,CARLSON S,et al.Reversible middle cerebral artery occlusion without craniectomy in rats[J].Stroke,1989,20(1):84-91.
[11]华兴邦,李辞蓉,周浩良,等.大鼠穴位图谱的研制[J].实验动物与动物实验,1991,7(1):1-5.
[12]闵友江,姚海华,邵水金,等.浅析《头针穴名国际标准化方案》的科学性[J].中国针灸,2007,27(8):612-616.
[13]徐叔云,卞如濂,陈修.药理实验方法学[M].3版.北京:人民卫生出版社,2002.
[14]SCHBITZ W R,BERGER C,KOLLMAR R,et al.Effect of brain-derived neurotrophic factor treatment and forced arm use on functional motor recovery after small cortical ischemia[J].Stroke,2004,35(4):992-997.
[15]ZGAVC T,CEULEMANS A G,HACHIMI-IDRISSI S,et al.The neuroprotective effect of post ischemic brief mild hypothermic treatment correlates with apoptosis,but not with gliosis in endothelin-1treated rats[J].BMC Neurosci,2012,13:105.
[16]CEULEMANS A G,ZGAVC T,KOOIJMAN R,et al.Mild hypothermia causes differential,time-dependent changes in cytokine expression and gliosis following endothelin-1-induced transient focal cerebral ischemia[J].J Neuroinflammation,2011,8:60.
[17]DI NAPOLI M,SHAH I M.Neuroinflammation and cerebrovascular disease in old age:a translational medicine perspective[J].J Aging Res,2011,2011:857484.
[18]LIU F D,MCCULLOUGH L D.Inflammatory responses in hypoxic ischemic encephalopathy[J].Acta Pharmacol Sin,2013,34(9):1121-1130.
[19]ORION D,SCHWAMMENTHAL Y,RESHEF T,et al.Interleukin-6and soluble intercellular adhesion molecule-1in acute brain ischaemia[J].Eur J Neurol,2008,15(4):323-328.
[20]PRADILLO J M,DENES A,GREENHALGH A D,et al.Delayed administration of interleukin-1receptor antagonist reduces ischemic brain damage and inflammation in comorbid rats[J].J Cereb Blood Flow Metab,2012,32(9):1810-1819.
[21]CASO J R,MORO M A,LORENZO P,et al.Involvement of IL-1βin acute stress-induced worsening of cerebral ischaemia in rats[J].Eur Neuropsychopharmacol,2007,17(9):600-607.
[22]SRTLE K,ZHOU J H,SHEN W H,et al.Interleukin-10in the brain[J].Crit Rev Immunol,2001,21(5):427-449.
[23]GRILLI M,BARBIERI I,BASUDEV H,et al.Interleukin-10modulates neuronal threshold of vulnerability to ischaemic damage[J].Eur J Neurosci,2000,12(7):2265-2272.
[24]GARCIA J M,STILLINGS S A,LECLERC J L,et al.Role of interleukin-10in acute brain injuries[J].Front Neurol,2017,8:244.
[25]VAN EXEL E,GUSSEKLOO J,DE CRAEN A J,et al.Inflammation and stroke:the Leiden 85-plus study[J].Stroke,2002,33(4):1135-1138.
[26]PREZ-DE PUIG I,MIRF,SALAS-PERDOMO A,et al.IL-10deficiency exacerbates the brain inflammatory response to permanent ischemia without preventing resolution of the lesion[J].J Cereb Blood Flow Metab,2013,33(12):1955-1966.
[27]HUXFORD T,GHOSH G.A structural guide to proteins of the NF-κB signaling module[J].Cold Spring Harb Perspect Biol,2009,1(3):a000075.
[28]DRIESSLER F,VENSTROM K,SABAT R,et al.Molecular mechanisms of interleukin-10-mediated inhibition of NF-κB activity:a role for p50[J].Clin Exp Immunol,2004,135(1):64-73.
[29]PFEILSCHIFTER W,CZECH B,HOFFMANN B P,et al.Pyrrolidine dithiocarbamate activates p38MAPK and protects brain endothelial cells from apoptosis:a mechanism for the protective effect in stroke[J].Neurochem Res,2010,35(9):1391-1401.
[30]WANG Z X,SHAN W R,CAO J B,et al.Early administration of pyrrolidine dithiocarbamate extends the therapeutic time window of tissue plasminogen activator in a male rat model of embolic stroke[J].J Neurosci Res,2018,96(3):449-458.