Please wait a minute...
浙江大学学报（医学版）
浙江大学学报（医学版）  2016, Vol. 45 Issue (5): 493-500    DOI: 10.3785/j.issn.1008-9292.2016.09.07
中药分子药理专题     
雷公藤甲素缓解局灶性脑组织缺血再灌注损伤诱发的炎症及其相关凋亡的研究
白石1,2, 孙雅谊2, 吴丽娟1, 吴仲敏1, 方马荣2
1. 台州学院医学院, 浙江 台州 318000;
2. 浙江大学医学院神经科学研究所, 浙江 杭州 310058
Tripotolide ameliorates inflammation and apoptosis induced by focal cerebral ischemia/reperfusion in rats
BAI Shi1,2, SUN Yayi2, WU Lijuan1, WU Zhongmin1, FANG Marong2
1. School of Medicine, Taizhou University, Taizhou 318000, China;
2. Institute of Neuroscience, Zhejiang University School of Medicine, Hangzhou 310058, China
 全文: PDF(16224 KB)   HTML ( ) 
摘要：

目的：探讨雷公藤甲素对局灶性脑组织缺血再灌注损伤大鼠脑组织的抗炎、抗凋亡等神经保护机制。方法：参照Longa的方法建立大鼠局灶性脑组织缺血再灌注损伤模型，80只SD大鼠随机分为健康对照组、手术对照组、溶剂对照组、模型对照组、雷公藤甲素组。TTC染色确定脑梗死的部位和体积，Longa法对神经功能障碍程度进行评分，荧光染色观察星形胶质细胞数量，TUNEL染色检测神经元凋亡情况，免疫组织化学检测诱导型一氧化氮合酶（iNOS）、环氧化酶-2（COX-2）、NF-κB的蛋白表达情况，实时定量PCR测定各组大鼠iNOS和COX-2 mRNA表达量的变化。结果：相较于溶剂对照组和模型对照组，雷公藤甲素组大鼠大脑中动脉分布区域脑组织水肿改善明显（80.03±0.46）%（P<0.05），诱发梗死体积缩小（8.3±1.4）%（P<0.01），神经功能障碍程度较轻，Longa评分为1.38±0.20（P<0.05）；炎症相关因素如GFAP阳性星形胶质细胞的数量减少（P<0.05），COX-2、iNOS和NF-κB蛋白表达下降，分别为91.67±1.31、95.24±5.07和75.03±12.06（均P<0.05），且损伤区凋亡作用减轻，TUNEL阳性细胞数为64.15±3.52（P<0.05）。结论：在大鼠局灶性脑组织缺血再灌注模型中，雷公藤甲素能够对抗炎症及其相关凋亡，起到减轻脑水肿及梗死范围，缓解神经功能性障碍的保护作用。
关键词： 雷公藤内酯/治疗应用;  脑组织缺血/药物疗法;  再灌注损伤/药物疗法;  神经炎/药物疗法;  NF-&kappa;  B/代谢;  环氧化酶2/代谢;  一氧化氮合酶/代谢;  疾病模型;  动物    
Abstract:

Objective: To investigate the effects of triptolide on inflammation and apoptosis induced by focal cerebral ischemia/reperfusion in rats. Methods: The rat model of focal cerebral ischemia/reperfusion injury was established according to Longa's method. A total of 80 SD rats were randomly divided into 5 groups:normal control, sham group, DMSO group, middle cerebral artery occlusion (MCAO) group, and MCAO with tripolide treatment group. TTC staining was used to examine the site and volume of cerebral infarction, and Longa score was employed for neurological disorders measurement. Number of astrocytes was measured by fluorescence staining, and neuronal apoptosis was determined by TUNEL staining. The expressions of inducible nitric oxide synthase(iNOS), cyclooxygenase 2(COX-2) and NF-κB proteins were detected by immunohistochemistry, and the expression of iNOS, COX-2 mRNA was detected by real-time PCR. Results: Compared with DMSO group and MCAO group, brain edema was improved (80.03±0.46)% (P<0.05), infarct volume was reduced (8.3±1.4)% (P<0.01), Longa score was decreased (1.38±0.20, P<0.05) in triptolide treatment group. Meanwhile triptolide also dramatically reduced the number of GFAP-positive astrocytes (P<0.05), alleviated protein expression of COX-2 (91.67±1.31), iNOS (95.24±5.07) and NF-κB (75.03±2.06) triggered by MCAO (all P<0.05), and induced a down-regulation of cell apoptosis as showed by TUNEL assay (64.15±3.52, P<0.05). Conclusion: Triptolide can reduce the cerebral infarction volume, attenuate brain edema and ameliorate the neurological deficits induced by cerebral ischemia-reperfusion injury rats, indicating that it might be used as a potential anti-inflammatory agent.
Key words: Triptolide/therapeutic use    Brain ischemia/drug therapy    Reperfusion injury/drug therapy    Neuritis/drug therapy    NF-kappa B/metabolism    Cyclooxygenase 2/metabolism    Nitric oxide synthase/metabolism    Disease models, animal
收稿日期: 2016-04-07 出版日期: 2016-09-25
CLC:  R743  
基金资助:

浙江省自然科学基金（LY15H090002）；国家自然科学基金（81671138）；浙江省科技厅公益技术应用研究（2015C37081，2013C37018）
通讯作者: 方马荣(1964-),男,博士,教授,博士生导师,从事神经科学研究;E-mail:fangmaro@zju.edu.cn;http://orcid.org/0000-0002-6636-4347     E-mail: fangmaro@zju.edu.cn
作者简介: 白石(1975-)男,硕士,讲师,从事神经生物学研究;E-mail:gaoxing5188@163.com;http://orcid.org/0000-0002-0746-3054
服务　　
把本文推荐给朋友
加入引用管理器
E-mail Alert
RSS
作者相关文章　　

引用本文:

白石 等. 雷公藤甲素缓解局灶性脑组织缺血再灌注损伤诱发的炎症及其相关凋亡的研究[J]. 浙江大学学报（医学版）, 2016, 45(5): 493-500.

BAI Shi, SUN Yayi, WU Lijuan, WU Zhongmin, FANG Marong. Tripotolide ameliorates inflammation and apoptosis induced by focal cerebral ischemia/reperfusion in rats. Journal of ZheJiang University(Medical Science), 2016, 45(5): 493-500.

链接本文:

http://www.zjujournals.com/med/CN/10.3785/j.issn.1008-9292.2016.09.07    或    http://www.zjujournals.com/med/CN/Y2016/V45/I5/493

[1] BAI S, HU Z, YANG Y, et al.Anti-inflammatory and neuroprotective effects of triptolide via the NF-κB signaling pathway in a rat MCAO model[J].Anat Rec (Hoboken), 2016, 299(2):256-266.
[2] XU Z, XU R X, LIU B S, et al.Time window characteristics of cultured rat hippocampal rat hippocampal neurons subjected to ischemia and reperfusion[J].Chin J Traumatol, 2005, 8(3):179-182.
[3] CHAN P H.Reactive oxygen radicals in signaling and damage in the ischemie brain[J].J Cereb Blood Flow Metab, 2001, 21(1):2-14.
[4] ZHAO G, VASZAR L T, QIU D, et al.Anti-inflammatory effects of triptolide in human bronchial epithelial cells[J].Am J Physiol Lung Cell Mol Physiol, 2000, 279(5):L958-L966.
[5] WANG X, LIANG X B, Li F Q, et al.Therapeutic strategies for Parkinson's disease:the ancient meets the future-traditional Chinese herbal medicine, electroacupuncture, gene therapy and stem cells[J].Neurochem Res, 2008, 33(10):1956-1963.
[6] GENG Y, FANG M, WANG J, et al.Triptolide down-regulates COX-2 expression and PGE2 release by suppressing the activity of NF-κB and MAP kinases in lipopolysaccharide-treated PC12 cells[J].Phytother Res, 2012, 26(3):337-343.
[7] XUE B, JIAO J, ZHANG L, et al.Triptolide upregulates NGF synthesis in rat astrocyte cultures[J].Neurochem Res, 2007, 32(7):1113-1119.
[8] SU Z, YUAN Y, CAO L, et al.Triptolide promotes spinal cord repair by inhibiting astrogliosis and inflammation[J].Glia, 2010, 58(8):901-915.
[9] LIU Y, CHEN H L, YANG G.Extract of Tripterygium wilfordii Hook F protect dopaminergic neurons against lipopolysaccharide-induced inflammatory damage[J].Am J Chin Med, 2010, 38(4):801-814.
[10] GUAN T, LIU Q, QIAN Y, et al.Ruscogenin reduces cerebral ischemic injury via NF-κB-mediated inflammatory pathway in the mouse model of experimental stroke[J].Eur J Pharmacol, 2013, 714(1-3):303-311.
[11] 陈卫伟, 杨留才, 潘施文, 等.线栓法制备SD大鼠局灶性脑缺血再灌注损伤模型[J].中国组织工程研究与临床康复, 2011, 15(50):9377-9380.CHEN Weiwei, YANG Liucai, PAN Shiwen, et al.Preparation of Sprague-Dawley rat models of focal cerebral ischemia-reperfusion injury by suture method[J].Chinese Journal of Tissue Engineering Research, 2011, 15(50):9377-9380.(in Chinese)
[12] 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.
[13] MDZINARISHVILI A, KIEWERT C, KUMAR V, et al.Bilobalide prevents ischemia-induced edema formation in vitro and in vivo[J].Neuroscience, 2007, 144(1):217-222.
[14] YANG F, LIU Y, TU J, et al.Activated astrocytes enhance the dopaminergic differentiation of stem cells and promote brain repair through bFGF[J/OL].Nat Commun, 2014, 5:5627.
[15] 李琳, 张志强.脑缺血再灌注损伤中细胞凋亡的研究进展[J].中华物理医学与康复杂志, 2005, 27(1):60-62.LI Lin, ZHANG Zhiqiang.Research progress of apoptosis in ischemia-reperfusion injury[J].Chinese Journal of Physical Medicine and Rehabiltation, 2005, 27(1):60-62.
[16] BRESGEN N, KARLHUBER G, KRIZBAI I, et al.Oxidative stress in cultured cerebral endothelial cells induces chromosomal aberrations, micronuclei, and apoptosis[J].J Neurosci Res, 2003, 72:327-333.
[17] DOHMEN C, KUMURA E, ROSNER G, et al.Extracellular correlates of glutamate toxicity in short-term cerebral ischemia and reperfusion:a direct in vivo comparison between white and gray matter[J].Brain Res, 2005, 1037(1-2):43-51.
[18] WEI J, QUAST M.Effect of nitric oxide synthesis inhibitor on a hyperglycemia rat model of reversible focal ischemia:detection of excitatory amino acids release and hydroxyl radical formation[J].Brain Res, 1998, 791(1-2):146-156.
[19] CEULEMANS AO, ZGAVC T, KOOIJMAN R, et al.The dual role of the neuroinflammatory response after ischemic stroke:modulatory effects of hypothermia[J].J Neuroinflammation, 2010, 7:74.
[20] SHICHITA T, SAKAGUCHI R, SUZUKI M, et al.Post-ischemic inflammation in the brain[J].Front Immunol, 2012, 3(3):132.
[21] DENES A, THORNTON P, ROTHWELL N J, et al.Inflammation and brain injury:acute cerebral ischaemia, peripheral and central inflammation[J].Brain Behav Immun, 2010, 24(5):708-723.
[22] YE Z, WANG N, XIA P, et al.Delayed administration of parecoxib, a specific COX-2 inhibitor, attenuated postischemic neuronal apoptosis by phosphorylation Akt and GSK-3β[J].Neurochem Res, 2012, 37(2):321-329.
[23] CLEMENS J A, STEPHENSON D T, YIN T, et al.Drug-induced neuroprotection from global ischemia is associated with prevention of persistent but not transient activation of nuclear factor-kappaB in rats[J].Stroke, 1998, 29(3):677-682.
[24] YASUDA Y, SHIMODA T, UNO K, et al.Temporal and sequential changes of glial cells and cytokine expression during neuronal degeneration after transient global ischemia in rats[J].J Neuroinflammation, 2011, 8:70.
[25] QIU D, ZHAO G, AOKI Y, et al.Immunosuppressant PG490(Triptolide) inhibits T-cell interleukin-2 expression at the level of purine-box/nuclear factor of activated T-cells and NF-kappaB transcriptional activation[J].J Biol Chem, 1999, 274(19):13443-13450.
[1] 曹洋,冯亚东,焦春花,施瑞华. 分段式可降解食管支架的临床前评价[J]. 浙江大学学报（医学版）, 2017, 46(6): 649-655.
[2] 张斌斌 等. 抑制哺乳动物雷帕霉素靶蛋白信号通路对慢性脑缺血小鼠认知功能的改善和机制[J]. 浙江大学学报（医学版）, 2017, 46(4): 405-412.
[3] 邹继霞,章程燕,王苹莉. 肿瘤过继T细胞免疫治疗应用进展[J]. 浙江大学学报（医学版）, 2017, 46(2): 211-217.
[4] 王莉,王瑜,武海英. 尼克酰胺降低妊娠期糖尿病大鼠的血糖水平以及调控线粒体超氧水平研究[J]. 浙江大学学报（医学版）, 2017, 46(2): 179-185.
[5] 王晏鹏,黄琼晓,徐盛,舒静. 采用吸宫和电凝方法建立小鼠子宫内膜损伤模型的比较[J]. 浙江大学学报（医学版）, 2017, 46(2): 186-191.
[6] 张力三,陈冠峰,陈洁芳,何旭东,胡兴越. 组胺改善癫痫所致记忆形成障碍的实验研究[J]. 浙江大学学报（医学版）, 2017, 46(1): 1-6.
[7] 高思倩,沈咏梅,耿福能,李艳华,高建青. 糖尿病溃疡动物模型的建立及相关治疗研究进展[J]. 浙江大学学报（医学版）, 2017, 46(1): 97-105.
[8] 陈立颖,汪仪,陈忠. 颞叶癫痫与海马成体神经再生[J]. 浙江大学学报（医学版）, 2017, 46(1): 22-29.
[9] 汪锡婷,唐藕凤,叶夷露,郑鸣之,胡珏,陈忠,钟恺. 藏红花素对小鼠电点燃癫痫的影响[J]. 浙江大学学报（医学版）, 2017, 46(1): 7-14.
[10] 侯甲福,武蕾蕾,才玉婷,高长久,安锦丹,于忠诚. 锦花清热胶囊抗内毒素作用实验研究[J]. 浙江大学学报（医学版）, 2017, 46(1): 74-79.
[11] 马婷婷,王毅,陈晓倩,赵筱萍. 液相色谱-质谱联用导向的黄葵胶囊肾保护活性物质研究[J]. 浙江大学学报（医学版）, 2017, 46(1): 66-73.
[12] 李统宇 等. 杜氏肌营养不良疾病模型及基因治疗研究进展[J]. 浙江大学学报（医学版）, 2016, 45(6): 648-654.
[13] 屈涛 等. 丹参素对去势大鼠骨质量的影响[J]. 浙江大学学报（医学版）, 2016, 45(6): 587-591.
[14] 方清清 等. 低频脉冲电磁场促进成骨细胞分化的基因调节和非基因调节探究[J]. 浙江大学学报（医学版）, 2016, 45(6): 568-574.
[15] 高玉海 等. 淫羊藿总黄酮胶囊对生长期大鼠骨密度和骨形态计量学的影响[J]. 浙江大学学报（医学版）, 2016, 45(6): 581-586.