补阳还五汤对实验性自身免疫性脑脊髓炎小鼠血脑屏障保护作用的研究
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摘要
目的:探讨补阳还五汤(BYHWD)对实验性自身免疫性脑脊髓炎(EAE)在发病潜伏期、高峰期和缓解期的血-脑屏障(BBB)保护的机制。方法:采用髓鞘MOG35-55诱导C57BL/6雌性小鼠建立EAE模型,随机分为EAE组和BYHWD组。于免疫后第3天开始灌胃给药,EAE组给予生理盐水,BYHWD组给予BYHWD,在免疫后的第9天、第17天和第28天,分别采集小鼠脑和脊髓标本,行HE染色和固蓝染色;采用Western Blot法检测脊髓和脑内细胞紧密连接蛋白Occludin、ZO-1的表达。结果:补阳还五汤治疗后可抑制中枢神经系统炎细胞浸润,减轻髓鞘脱失;与EAE对照组比较,BYHWD治疗9 d、17 d和28 d,脊髓和脑内紧密连接蛋白Occludi和ZO-1表达增加。结论:补阳还五汤可以通过上调脊髓和脑内细胞紧密连接蛋白Occludin、ZO-1的表达,发挥血脑屏障(BBB)完整性的保护,从而发挥神经保护的作用。
Objective:To investigate the mechanism of Buyang Huanwu decoction(BYHWD)in the protection of blood brain barrier(BBB)in experimental autoimmune encephalomyelitis(EAE)during the incubation period,peak period and remediation period. Methods:EAE model was established in female C57BL/6 mice induced by myelin sheath Mog35-55 and randomly divided into EAE group and BYHWD group. The EAE group was perfused normal saline and BYHWD group was perfused BYHWD on the 3~(rd) day after immunization.Brain and spinal cord specimens of mice were collected on the 9~(th) day,17~(th) day and 28~(th) day after immunization,respectively,and HE staining and Luxol Fast Blue were performed.Western blot was used to detect the expression of Occludin and ZO-1 in spinal cord and brain cells. Results:Buyang Huanwu decoction can inhibit the infiltration of inflammatory cells in central nervous system and alleviate the loss of myelin sheath after treatment. Compared with EAE group,BYHWD treatment in BYHWD group the expression of occludin and ZO-1 at 9 days,17 days and 28 days were increased in the spinal cord and brain. Conclusion:Buyang Huanwu decoction can play a neuroprotective role by up-regulating the expression of tight junction protein Occludin and ZO-1 in spinal cord and brain cells,so as to protect the integrity of BBB.
Objective:To investigate the mechanism of Buyang Huanwu decoction(BYHWD)in the protection of blood brain barrier(BBB)in experimental autoimmune encephalomyelitis(EAE)during the incubation period,peak period and remediation period. Methods:EAE model was established in female C57BL/6 mice induced by myelin sheath Mog35-55 and randomly divided into EAE group and BYHWD group. The EAE group was perfused normal saline and BYHWD group was perfused BYHWD on the 3~(rd) day after immunization.Brain and spinal cord specimens of mice were collected on the 9~(th) day,17~(th) day and 28~(th) day after immunization,respectively,and HE staining and Luxol Fast Blue were performed.Western blot was used to detect the expression of Occludin and ZO-1 in spinal cord and brain cells. Results:Buyang Huanwu decoction can inhibit the infiltration of inflammatory cells in central nervous system and alleviate the loss of myelin sheath after treatment. Compared with EAE group,BYHWD treatment in BYHWD group the expression of occludin and ZO-1 at 9 days,17 days and 28 days were increased in the spinal cord and brain. Conclusion:Buyang Huanwu decoction can play a neuroprotective role by up-regulating the expression of tight junction protein Occludin and ZO-1 in spinal cord and brain cells,so as to protect the integrity of BBB.
引文
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[15]Melanie D Sweeney,Abhay P Sagare,Berislav V.Zlokovic.Blood-brain barrier breakdown in Alzheimer′s disease and other neurodegenerative disorders[J].Nat Rev Neurol,2018,14(3):133-150.
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[17]李丹丹,胡景霞,白波.CD4+T细胞和小胶质细胞在实验性自身免疫性脑脊髓炎中的作用[J].现代免疫学,2012,32(3):261-264.
[18]葛建彬,宋新建,陈丹丹,等.紫菜多糖对脑缺血再灌注损伤小鼠血脑屏障的保护作用[J].中国临床药理学杂志,34(19):2291-2294.
[19]Bennet J,Basivireddy J,Kollar A,et al.Blood brain barrier disruption and enhanced vascular permeability in the multiple sclerosis model EAE[J].Neuroimmunol,2010,229(1/2):180-191.
[20]Kooij G,Mizee MR,van HJ,et al.Adenosine riphosphate-binding cassette transporters mediate chemokine(C-C motif)ligand 2 secretion from reactive astrocytes:relevance to multiple sclerosis pathogenesis[J].Brain,2011(134):555-570.
[21]Kirk J,Plumb J,Mirakhur M,et al.Tight junctional abnormality in multiple sclerosis white matter affects all calibres of vessel and is associated with blood-brain barrier leakage and active demyelination[J].J Pathol,2003(201):319-327.
[2]于婧文,李艳花,张辉,等.新型Rho激酶抑制剂FSD-C11化合物抑制EAE的作用研究[J].中国神经免疫学和神经病学杂志,2015,22(3):156-161.
[3]吴彦青,张玉莲,高颖,等.当代医家论治多发性硬化经验探析[J].中华中医药学刊,2013,31(8):1781-1784.
[4]周哲屹,卢昌均,汪鸿浩,等.中医药治疗多发性硬化的研究进展[J].山东中医杂志,2017,36(1):74-77.
[5]吴畏.多发性硬化中医证候学规律的研究[D].北京:北京中医药大学,2013.
[6]王晓庆,田倩倩,李艳花,等.补阳还五汤对EAE小鼠免疫调节的作用[J].中国免疫学杂志,2017,33(1):52-57.
[7]田倩倩,李艳花,尉杰忠,等.补阳还五汤对实验性自身免疫性脑脊髓炎单核巨噬细胞的免疫调控作用[J].中国病理生理杂志,2017,33(2):200-207.
[8]田倩倩.补阳还五汤对实验性自身免疫性脑脊髓炎单核巨噬细胞的免疫调控研究[D].太原:山西中医学院,2017.
[9]Wang Xiaqing,Yang Chunyan,Li Yanhua,et al.Immunoregulative effect of Buyang Huanwu Decoction in experimental autoimmune encephalomyelitis[J].Int J Clin Exp Med,2017,10(3):129-131.
[10]谷青芳,张辉,李艳花,等.法舒地尔对EAE小鼠血-脑屏障功能的保护作用及机制研究[J].中国神经免疫学和神经病学杂志,2017,24(2):99-104.
[11]Lou ZY,Chen C,He Q,et al.Targeting CB(2)receptor as a neuro inflammatory modulator in experimental autoimmune encephalomyelitis[J].Mol lmmunol,2011,49(3):453-461.
[12]Urban JL,Kumar V,Kono DH,et al.Restricted use of T cell receptor V genes in murine autoimmune encephalomyelitis raises possibilities for antibody therapy[J].Cell,1988,54(4):577-592.
[13]周忠光.组织学与胚胎学[M].北京:中国中医药出版社,2016.
[14]Tan F,Fuw J,Cheng NF,et al.Ligustrazine reduces blood-brain barrier permeability in a rat model of focal cerebral ischemia and reperfusion[J].Exp Ther Med,2015,9(5):1757-1762.
[15]Melanie D Sweeney,Abhay P Sagare,Berislav V.Zlokovic.Blood-brain barrier breakdown in Alzheimer′s disease and other neurodegenerative disorders[J].Nat Rev Neurol,2018,14(3):133-150.
[16]Engelhardt B.Vascular,glial,and lymphatic immune gateways of the central nervous system[J].Acta Neuropathol(Berl),2016(132):317-338.
[17]李丹丹,胡景霞,白波.CD4+T细胞和小胶质细胞在实验性自身免疫性脑脊髓炎中的作用[J].现代免疫学,2012,32(3):261-264.
[18]葛建彬,宋新建,陈丹丹,等.紫菜多糖对脑缺血再灌注损伤小鼠血脑屏障的保护作用[J].中国临床药理学杂志,34(19):2291-2294.
[19]Bennet J,Basivireddy J,Kollar A,et al.Blood brain barrier disruption and enhanced vascular permeability in the multiple sclerosis model EAE[J].Neuroimmunol,2010,229(1/2):180-191.
[20]Kooij G,Mizee MR,van HJ,et al.Adenosine riphosphate-binding cassette transporters mediate chemokine(C-C motif)ligand 2 secretion from reactive astrocytes:relevance to multiple sclerosis pathogenesis[J].Brain,2011(134):555-570.
[21]Kirk J,Plumb J,Mirakhur M,et al.Tight junctional abnormality in multiple sclerosis white matter affects all calibres of vessel and is associated with blood-brain barrier leakage and active demyelination[J].J Pathol,2003(201):319-327.
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