法舒地尔保护小鼠脑缺血再灌流损伤的机制
|
摘要
目的探讨法舒地尔(Fasudil)保护小鼠脑缺血/再灌流(ischemia/reperfusion,I/R)损伤的作用机制。方法对成年C57BL/6J雄性小鼠进行法舒地尔灌胃处理,然后行大脑中动脉闭塞(middle cerebral artery occlusion,MCAo)动物模型,用氯化三苯四氮唑染色法(2,3,5-triphenyltetrazolium chloride,TTC)染色以分析脑梗死情况,激光散斑(Laser Speckle)图像系统分析脑血流改变情况;对成年C57BL/6J雄性小鼠进行法舒地尔灌胃处理后,取脑组织用免疫印迹分析过氧化物酶体激活受体α(peroxisome proliferators-activated receptor alpha,PPARα)的表达状态,实时荧光定量PCR检测超氧化物歧化酶(superoxide dismutase,SOD)1～3的表达及化学发光法检测SODs的活性。结果雄性C57BL/6J小鼠经法舒地尔处理后,脑组织中PPARα和SOD1、SOD2、SOD3的表达升高(P=0.000、0.000、0.000、0.000),SODs酶活性升高(P=0.000);在小鼠脑MCAO动物模型中,法舒地尔预处理使脑坏死体积明显减小(P=0.000),再灌流后脑坏死区域的脑血流强于羟甲基纤维素(carboxymethyl cellulose,CMC)处理小鼠。结论法舒地尔通过促进PPARα的表达,影响SOD1、SOD2、SOD3的表达及活性,以改善小鼠脑缺血区域的脑血流,从而保护I/R损伤。
Objective To investigate the neuroprotective mechanisms of Fasudil on brain ischemia/reperfusion injury with MCAO model. Methods Separate C57 BL/6 J mice was treated with Fasudil(10 mg/kg.BW) through intragastric administration for 3 days, while the mice treated with CMC(10 mL 0.5%CMC/kg.BW) were used as control group. The treated mice were subjected to 60 minutes of focal ischemia and 18 hours reperfusion. The brain was collected and the infarct volume was analyzed through TTC staining with MCID image system. Cerebral blood flow was measured with laser speckle in MCAO. The brain collected from the mice treated with Fasudil or CMC, without MCAO, was used to check the expression of PPAR-alpha, SOD1, 2, 3 and the activity of SODs. The expression of PPAR-alpha was checked with immuno-blotting assay, the mRNA of SOD1, 2, 3 was assayed with real-time PCR, and the activity of SODs was analyzed with a chemiluminescence method. Results Compared with control group, the PPAR-alpha and SOD1, 2, 3 in the brain from Fasudil treated mice was significantly higher(P=0.000). The activity of SODs in the Fasudil treated mice was significantly higher than control group(P=0.000). The infarct volume was significantly attenuated with application of Fasudil(P=0.000). Compared with the control group, the cerebral blood flow was attenuated with application of Fasudil. Conclusion The neuroprotective mechanisms of Fasudil on the brain ischemia/reperfusion injury is through promoting the expression of PPAR-alpha to boost the expression of SOD1, 2, 3 and the activity of SODs to enhance the cerebral blood flow to attenuate the I/R injury.
Objective To investigate the neuroprotective mechanisms of Fasudil on brain ischemia/reperfusion injury with MCAO model. Methods Separate C57 BL/6 J mice was treated with Fasudil(10 mg/kg.BW) through intragastric administration for 3 days, while the mice treated with CMC(10 mL 0.5%CMC/kg.BW) were used as control group. The treated mice were subjected to 60 minutes of focal ischemia and 18 hours reperfusion. The brain was collected and the infarct volume was analyzed through TTC staining with MCID image system. Cerebral blood flow was measured with laser speckle in MCAO. The brain collected from the mice treated with Fasudil or CMC, without MCAO, was used to check the expression of PPAR-alpha, SOD1, 2, 3 and the activity of SODs. The expression of PPAR-alpha was checked with immuno-blotting assay, the mRNA of SOD1, 2, 3 was assayed with real-time PCR, and the activity of SODs was analyzed with a chemiluminescence method. Results Compared with control group, the PPAR-alpha and SOD1, 2, 3 in the brain from Fasudil treated mice was significantly higher(P=0.000). The activity of SODs in the Fasudil treated mice was significantly higher than control group(P=0.000). The infarct volume was significantly attenuated with application of Fasudil(P=0.000). Compared with the control group, the cerebral blood flow was attenuated with application of Fasudil. Conclusion The neuroprotective mechanisms of Fasudil on the brain ischemia/reperfusion injury is through promoting the expression of PPAR-alpha to boost the expression of SOD1, 2, 3 and the activity of SODs to enhance the cerebral blood flow to attenuate the I/R injury.
引文
[1] Favate AS,Younger DS.Epidemiology of Ischemic Stroke[J].Neurol Clin,2016,34(4):967-980.
[2] Sun J,Guo Y,Wang X,et al.mHealth For Aging China:Opportunities and Challenges[J].Aging Dis,2016,7(1):53-67.
[3] 刘星芳,张秀梅,单热爱.脑缺血/再灌注损伤对血脑屏障的影响及干预措施的研究进展[J].广东医学,2016,37(21):3292-3294.
[4] Zhang HZ,Jiang W,Zhao XF,et al.Electroacupuncture induces acute changes in cerebral cortical miRNA profile,improves cerebral blood flow and alleviates neurological deficits in a rat model of stroke[J].Neural Regen Res,2016,11(12):1940-1950.
[5] Fukuta T,Asai T,Sato A,et al.Neuroprotection against cerebral ischemia/reperfusion injury by intravenous administration of liposomal fasudil[J].Int J Pharm,2016,506(1/2):129-137.
[6] Paintlia AS,Paintlia MK,Singh AK,et al.Modulation of Rho-Rock signalingpathway protects oligodendrocytes against cytokine toxicity via PPAR-alpha-dependent mechanism[J].Glia,2013,61(9):1500-1517.
[7] Yang Y,Rong X,Lv X,et al.Inhibition of mevalonate pathway prevents ischemia-induced cardiac dysfunction in rats via RhoA-independent signaling pathway[J].Cardiovasc Ther,2017,35(5):1-9.
[8] FuKuta T,Asai T,Yanagida Y,et al.Combination therapy with liposomal neuroprotectants and tissue plasminogen activator for treatment of ischemic stroke[J].FASEB J,2017,31(5):1879-1890.
[9] Ohata K,Chen-Yoshikawa TF,Menju T,et al.Protective Effect of Inhaled Rho-Kinase Inhibitor on Lung Ischemia-Reperfusion Injury[J].Ann Thorac Surg,2017,103(2):476-483.
[10] 李继红,杨锡彤,杜小珊,等.非诺贝特调控SOD1表达的机制[J].广东医学,2018,39(1):68-75.
[11] Shen L,Zhang P,Zhang S,et al.C-X-C motif chemokine ligand 8 promotes endothelial cell homing via the Akt-signal transducer and activator of transcription pathway to accelerate healing of ischemic and hypoxic skin ulcers[J].Exp Ther Med,2017,13(6):3021-3031.
[12] Deska M,Romuk E,Segiet OA,et al.Oxidative stress and angiogenesis in primary hyperparathyroidism[J].Eur Surg,2017,49(3):118-126.
[13] Han T,Liu M,Yang S.DJ-1 Alleviates Angiotensin II-Induced Endothelial Progenitor Cell Damage by Activating the PPARγ/HO-1 Pathway[J].Cell Biochem,2018,119(1):392-400.
[14] 程建杰,杨锡彤,杜小珊,等.法舒地尔对小鼠脑缺血再灌注损伤保护作用的血脑屏障机制[J].中华行为医学与脑科学杂志,2017,26(10):933-937.
[15] Wang G,Liu X,Guo Q,et al.Chronic treatment with fibrates elevates superoxide dismutase in adult mouse brain microvessels[J].Brain Res,2010,1359:247-255.
[16] Guo Q,Wang G,Namura S.Fenofibrate improves cerebral blood flow after middle cerebral artery occlusion in mice[J].Cereb Blood Flow Metab,2010,30(1):70-78.
[17] Wang G,Guo Q,Hossain M,et al.Bone marrow-derived cells are the major source of MMP-9 contributing to blood-brain barrier dysfunction and infarct formation after ischemic stroke in mice[J].Brain Res,2009,1294:183-192.
[2] Sun J,Guo Y,Wang X,et al.mHealth For Aging China:Opportunities and Challenges[J].Aging Dis,2016,7(1):53-67.
[3] 刘星芳,张秀梅,单热爱.脑缺血/再灌注损伤对血脑屏障的影响及干预措施的研究进展[J].广东医学,2016,37(21):3292-3294.
[4] Zhang HZ,Jiang W,Zhao XF,et al.Electroacupuncture induces acute changes in cerebral cortical miRNA profile,improves cerebral blood flow and alleviates neurological deficits in a rat model of stroke[J].Neural Regen Res,2016,11(12):1940-1950.
[5] Fukuta T,Asai T,Sato A,et al.Neuroprotection against cerebral ischemia/reperfusion injury by intravenous administration of liposomal fasudil[J].Int J Pharm,2016,506(1/2):129-137.
[6] Paintlia AS,Paintlia MK,Singh AK,et al.Modulation of Rho-Rock signalingpathway protects oligodendrocytes against cytokine toxicity via PPAR-alpha-dependent mechanism[J].Glia,2013,61(9):1500-1517.
[7] Yang Y,Rong X,Lv X,et al.Inhibition of mevalonate pathway prevents ischemia-induced cardiac dysfunction in rats via RhoA-independent signaling pathway[J].Cardiovasc Ther,2017,35(5):1-9.
[8] FuKuta T,Asai T,Yanagida Y,et al.Combination therapy with liposomal neuroprotectants and tissue plasminogen activator for treatment of ischemic stroke[J].FASEB J,2017,31(5):1879-1890.
[9] Ohata K,Chen-Yoshikawa TF,Menju T,et al.Protective Effect of Inhaled Rho-Kinase Inhibitor on Lung Ischemia-Reperfusion Injury[J].Ann Thorac Surg,2017,103(2):476-483.
[10] 李继红,杨锡彤,杜小珊,等.非诺贝特调控SOD1表达的机制[J].广东医学,2018,39(1):68-75.
[11] Shen L,Zhang P,Zhang S,et al.C-X-C motif chemokine ligand 8 promotes endothelial cell homing via the Akt-signal transducer and activator of transcription pathway to accelerate healing of ischemic and hypoxic skin ulcers[J].Exp Ther Med,2017,13(6):3021-3031.
[12] Deska M,Romuk E,Segiet OA,et al.Oxidative stress and angiogenesis in primary hyperparathyroidism[J].Eur Surg,2017,49(3):118-126.
[13] Han T,Liu M,Yang S.DJ-1 Alleviates Angiotensin II-Induced Endothelial Progenitor Cell Damage by Activating the PPARγ/HO-1 Pathway[J].Cell Biochem,2018,119(1):392-400.
[14] 程建杰,杨锡彤,杜小珊,等.法舒地尔对小鼠脑缺血再灌注损伤保护作用的血脑屏障机制[J].中华行为医学与脑科学杂志,2017,26(10):933-937.
[15] Wang G,Liu X,Guo Q,et al.Chronic treatment with fibrates elevates superoxide dismutase in adult mouse brain microvessels[J].Brain Res,2010,1359:247-255.
[16] Guo Q,Wang G,Namura S.Fenofibrate improves cerebral blood flow after middle cerebral artery occlusion in mice[J].Cereb Blood Flow Metab,2010,30(1):70-78.
[17] Wang G,Guo Q,Hossain M,et al.Bone marrow-derived cells are the major source of MMP-9 contributing to blood-brain barrier dysfunction and infarct formation after ischemic stroke in mice[J].Brain Res,2009,1294:183-192.