温阳补肾灸对血管性痴呆大鼠NF-κB信号通路的影响（英文）
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摘要
目的:观察温阳补肾灸对血管性痴呆(VD)大鼠行为学、海马IL-1β、TNF-α及NF-κB相关基因与蛋白表达的影响,探讨温阳补肾灸抑制VD炎性反应的机制。方法:SD大鼠随机分为假手术组、模型组、艾灸组、西药组,每组10只。采用双侧颈总动脉缺血再灌注法制造VD大鼠模型。造模后,艾灸组大鼠悬灸"大椎""命门""关元",每次15 min,每日一次,连续治疗4周;西药组大鼠用尼莫地平灌胃(2 mg·kg~(-1)·d~(-1))连续治疗4周。运用Morris水迷宫测试对各组大鼠进行行为学检测。用HE染色观察其组织病理学改变。运用RT-qPCR法和Westernblot法检测海马IL-1β、TNF-α及NF-κB相关蛋白与基因的表达。结果:造模后手术组大鼠Morris水迷宫逃避潜伏期较假手术组明显延长(P<0.05)。HE染色显示模型组神经元细胞排列松散,细胞膜模糊,细胞质不均匀,核固缩,坏死量增多,炎细胞显著增加。与模型组相比,艾灸组与西药组炎细胞数目明显减少,神经元坏死减少。治疗后,与模型组比较,艾灸组、西药组大鼠海马组织TNF-α、p-IκB、NF-κBp65蛋白表达量显著降低(P<0.05);与模型组比较,艾灸组、西药组大鼠海马组织IL-1β、TNF-α、NF-κBp65 mRNA表达显著降低(P<0.05)。结论:温阳补肾灸可能通过降低脑内IL-1β、TNF-α及NF-κB相关基因的表达水平,抑制NF-κB信号通路,降低炎性反应,以达到治疗VD的目的。
Objective: To observe the effect of warming yang and supplementing kidney moxibustion therapy on the behavioristics and the expression levels of hippocampal IL-1β, TNF-α and NF-κB related genes and proteins of vascular dementia(VD) rats, and to explore the mechanism of warming yang and supplementing kidney moxibustion therapy in inhibiting VD inflammatory response. Methods Seventy-eight SD rats, except 10 rats as sham operation group(group A), were established into VD models by applying the ischemia reperfusion method in bilateral common carotid arteries. 30 models were successful and were randomly divided into model group(group B), moxibustion group(group C) and western medication group(group D), with 10 rats in each group. After modeling, suspended moxibustion was conducted at "Dàzhuī(大椎GV 14)","Mìngmén(命门GV 4)" and "Guānyuán(关元CV 4)" of the rats in group C for15 min/time. The intervention was performed for once a day, and intervention for 4 consecutive weeks was needed. Intragastric administration with nimodipine(2 mgkg~(-1)d~(-1)) was carried out in the rats in group D for 4 consecutive weeks. Morris water maze test was adopted for behavioral test in the rats in each group. HE staining was conducted in order to observe the pathological changes. RT-qPCR method and Western blot method were used for detecting the expression levels of hippocampal IL-1β, TNF-αand NF-κB related genes and proteins.Results: After modeling, the Morris water maze escape latency of rats in operation groups was obviously longer than that in group A(P<0.05). According to HE staining results, the arrangement of neurons in group B was incompact, the cell membranes were undefined, the cytoplasm was heterogeneous, karyopyknosis was found, the amount of necrosis increased, and the number of inflammatory cells significantly increased. Compared with group B, the number of inflammatory cells significantly reduced in group C and group D, and the amount of neuron necrosis also reduced. After intervention, compared with group B, the expression levels of TNF-a, p-IκB and NF-κBp65 mRNA in the hippocampal of the rats in group C and group D significantly decreased(all P<0.05), compared with group B, the expression levels of IL-1β, TNF-α and NF-κBp65 mRNA in the hippocampal of rats in group C and group D also significantly decreased(all P<0.05).Conclusion: Warming yang and supplementing kidney moxibustion therapy may inhibit the NF-κB signaling pathway and reduce the inflammatory response through lowering the expression levels of IL-1β,TNF-α and NF-κB mRNA in the brain, thus achieving the purpose of treating VD.
Objective: To observe the effect of warming yang and supplementing kidney moxibustion therapy on the behavioristics and the expression levels of hippocampal IL-1β, TNF-α and NF-κB related genes and proteins of vascular dementia(VD) rats, and to explore the mechanism of warming yang and supplementing kidney moxibustion therapy in inhibiting VD inflammatory response. Methods Seventy-eight SD rats, except 10 rats as sham operation group(group A), were established into VD models by applying the ischemia reperfusion method in bilateral common carotid arteries. 30 models were successful and were randomly divided into model group(group B), moxibustion group(group C) and western medication group(group D), with 10 rats in each group. After modeling, suspended moxibustion was conducted at "Dàzhuī(大椎GV 14)","Mìngmén(命门GV 4)" and "Guānyuán(关元CV 4)" of the rats in group C for15 min/time. The intervention was performed for once a day, and intervention for 4 consecutive weeks was needed. Intragastric administration with nimodipine(2 mgkg~(-1)d~(-1)) was carried out in the rats in group D for 4 consecutive weeks. Morris water maze test was adopted for behavioral test in the rats in each group. HE staining was conducted in order to observe the pathological changes. RT-qPCR method and Western blot method were used for detecting the expression levels of hippocampal IL-1β, TNF-αand NF-κB related genes and proteins.Results: After modeling, the Morris water maze escape latency of rats in operation groups was obviously longer than that in group A(P<0.05). According to HE staining results, the arrangement of neurons in group B was incompact, the cell membranes were undefined, the cytoplasm was heterogeneous, karyopyknosis was found, the amount of necrosis increased, and the number of inflammatory cells significantly increased. Compared with group B, the number of inflammatory cells significantly reduced in group C and group D, and the amount of neuron necrosis also reduced. After intervention, compared with group B, the expression levels of TNF-a, p-IκB and NF-κBp65 mRNA in the hippocampal of the rats in group C and group D significantly decreased(all P<0.05), compared with group B, the expression levels of IL-1β, TNF-α and NF-κBp65 mRNA in the hippocampal of rats in group C and group D also significantly decreased(all P<0.05).Conclusion: Warming yang and supplementing kidney moxibustion therapy may inhibit the NF-κB signaling pathway and reduce the inflammatory response through lowering the expression levels of IL-1β,TNF-α and NF-κB mRNA in the brain, thus achieving the purpose of treating VD.
引文
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[3]Jin R.Jin ZG,Cai H.Effect of acupuncture combined with geniposide on the microglial cell activation and the expression level of NF-κB in brain tissue of rats with diabetes mellitus combined with cerebral ischemia.Acup Res2014;39(4):305-12.
[4]Chen SH,Sun H,Xu H,Zhang YM,Gao Y,Li S.Effect of acupuncture on the expression of interleukin-1βand tumor necrosis factor-βin bilateral brain tissue of rats with cerebral ischemia reperfusion injury.Acup Res 2012:37(6):470-5.
[5]Jiang JF,Wang LL,Xu B,Ling H,Song XG.Wu HG.Antiinflammation-the effect mechanism of moxibustion.Zhongguo Zhen Jiu 2013:33(9):860-4.
[6]Zhang H.Shuang Z,Song X,Xu W,Li L.Su S.et al.Regulating effect of moxibustion on MCP-1 and NF-κB in the colonic mucosa tissue of rats with Crohn's disease.World J Acup-Mox 2016;26(4):33-40.
[7]Wang P.Yang J,Yang F,Chen H.Clinical observation of dissolving stasis and unblocking collaterals moxibustion therapy for treatment of vascular dementia.J Emerg Tradit Chin Med 2009;18(2):172-4.
[8]Zhu CF,Yang J,Yang K,Cai SZ,Fei AH,Yu HW,et al.Clinical observation of unblocking governor vessel and dissolving stasis moxibustion therapy for treatment of vascular cognitive disorder.Shanghai J Acup-Mox 2013:32(4):242-4.
[9]Sheng DD,Cai SZ.Efficacy observation of warming yang and supplementing kidney moxibustion therapy for treatment of vascular dementia.J Pract Tradit Chin Med 2017;33(1):58-9.
[10]Chen H.Wang P,Yang J,Liu G.Effect of moxibustion on the symptoms and the level of neuropeptides in the cerebrospinal fluid of patients with vascular dementia.World J Acup-Mox 2011;21(2):001.
[11]Wang P,Tang JY,Yang J,Zhang QP.Effect of moxibustion on the expression levels of hippocampal VEGF,flt-1,bFGF and bFGF-r of rats with vascular dementia.Chin J Integr Tradit West Med 2012;32(1):97-101.
[12]Lin YP.Wan QQ,Peng Y,He FE,Shen J.Effect of electro-acupuncture on the expression levels of somatotropin mRNA and growth hormone secretagogue receptor mRNA in the antrum of stomach of rats with diabetes mellitus and gastroparesis.Acup Res 2015;40(4):290-5.
[13]Rius J,Guma M.Schachtrup C,Akassoglou K,Zinkemagel AS,Nizet V,et al.NF-κB links innate immunity to the hypoxic response through transcriptional regulation of HIF-1α.Nature 2008;453(7196):807-11.
[14]Chen Y,Zhang L,Ni J,Wang X,Cheng J,Li Y,et al.LLDT-8 protects against cerebral ischemia/reperfusion injury by suppressing post-stroke inflammation.J Pharmacol Sci 2016:131(2):131-7.
[15]Ha SK,Moon E,Kim SY.Chrysin suppresses LPS-stimulated proinflammatory responses by blocking NF-κB and JNK activations in microglia cells.Neurosci Lett 2010;485(3):143-7.
[16]Qjn WY,Luo Y.Yu C.Effect of electro-acupuncture on the expression levels of hippocampal IL-1βand NF-κB of rats with focal cerebral ischemia reperfusion.Acup Res 2013;38(4):271-6.
[17]Zhu CF,Sun JJ.Han W,Yang J.Study of the mechanism of unblocking governor vessel and regulating the mind moxibustion therapy in the inhibition of over-expression of hippocampalβ-amyloid protein of rats with mild cognitive impairment.Acup Res 2016;41(2):131-7.
[18]Wang P,Yang J,Zhang QP,Shi HP,Ma L Effect of moxibustion at the head acupoints on the expression levels of VEGF and the receptors Flt-1 and Flk-1mRNA in brain tissue of VD rats.J Emerg Tradit Chin Med 2013;22(6):861-3.
[2]ladecola C.The pathobiology of vascular dementia.Neuron 2013;80(4):844-66.
[3]Jin R.Jin ZG,Cai H.Effect of acupuncture combined with geniposide on the microglial cell activation and the expression level of NF-κB in brain tissue of rats with diabetes mellitus combined with cerebral ischemia.Acup Res2014;39(4):305-12.
[4]Chen SH,Sun H,Xu H,Zhang YM,Gao Y,Li S.Effect of acupuncture on the expression of interleukin-1βand tumor necrosis factor-βin bilateral brain tissue of rats with cerebral ischemia reperfusion injury.Acup Res 2012:37(6):470-5.
[5]Jiang JF,Wang LL,Xu B,Ling H,Song XG.Wu HG.Antiinflammation-the effect mechanism of moxibustion.Zhongguo Zhen Jiu 2013:33(9):860-4.
[6]Zhang H.Shuang Z,Song X,Xu W,Li L.Su S.et al.Regulating effect of moxibustion on MCP-1 and NF-κB in the colonic mucosa tissue of rats with Crohn's disease.World J Acup-Mox 2016;26(4):33-40.
[7]Wang P.Yang J,Yang F,Chen H.Clinical observation of dissolving stasis and unblocking collaterals moxibustion therapy for treatment of vascular dementia.J Emerg Tradit Chin Med 2009;18(2):172-4.
[8]Zhu CF,Yang J,Yang K,Cai SZ,Fei AH,Yu HW,et al.Clinical observation of unblocking governor vessel and dissolving stasis moxibustion therapy for treatment of vascular cognitive disorder.Shanghai J Acup-Mox 2013:32(4):242-4.
[9]Sheng DD,Cai SZ.Efficacy observation of warming yang and supplementing kidney moxibustion therapy for treatment of vascular dementia.J Pract Tradit Chin Med 2017;33(1):58-9.
[10]Chen H.Wang P,Yang J,Liu G.Effect of moxibustion on the symptoms and the level of neuropeptides in the cerebrospinal fluid of patients with vascular dementia.World J Acup-Mox 2011;21(2):001.
[11]Wang P,Tang JY,Yang J,Zhang QP.Effect of moxibustion on the expression levels of hippocampal VEGF,flt-1,bFGF and bFGF-r of rats with vascular dementia.Chin J Integr Tradit West Med 2012;32(1):97-101.
[12]Lin YP.Wan QQ,Peng Y,He FE,Shen J.Effect of electro-acupuncture on the expression levels of somatotropin mRNA and growth hormone secretagogue receptor mRNA in the antrum of stomach of rats with diabetes mellitus and gastroparesis.Acup Res 2015;40(4):290-5.
[13]Rius J,Guma M.Schachtrup C,Akassoglou K,Zinkemagel AS,Nizet V,et al.NF-κB links innate immunity to the hypoxic response through transcriptional regulation of HIF-1α.Nature 2008;453(7196):807-11.
[14]Chen Y,Zhang L,Ni J,Wang X,Cheng J,Li Y,et al.LLDT-8 protects against cerebral ischemia/reperfusion injury by suppressing post-stroke inflammation.J Pharmacol Sci 2016:131(2):131-7.
[15]Ha SK,Moon E,Kim SY.Chrysin suppresses LPS-stimulated proinflammatory responses by blocking NF-κB and JNK activations in microglia cells.Neurosci Lett 2010;485(3):143-7.
[16]Qjn WY,Luo Y.Yu C.Effect of electro-acupuncture on the expression levels of hippocampal IL-1βand NF-κB of rats with focal cerebral ischemia reperfusion.Acup Res 2013;38(4):271-6.
[17]Zhu CF,Sun JJ.Han W,Yang J.Study of the mechanism of unblocking governor vessel and regulating the mind moxibustion therapy in the inhibition of over-expression of hippocampalβ-amyloid protein of rats with mild cognitive impairment.Acup Res 2016;41(2):131-7.
[18]Wang P,Yang J,Zhang QP,Shi HP,Ma L Effect of moxibustion at the head acupoints on the expression levels of VEGF and the receptors Flt-1 and Flk-1mRNA in brain tissue of VD rats.J Emerg Tradit Chin Med 2013;22(6):861-3.
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