氢饱和生理盐水治疗大鼠脑缺血再灌注损伤的效果研究
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摘要
目的:探讨氢饱和生理盐水对大鼠脑缺血再灌注损伤模型的治疗效果及可能的作用机制。方法:SD大鼠(250-300g)随机分为3组(n=20):假手术组(Sham组),缺血再灌注损伤模型组(I/R组),氢饱和盐水治疗组(HS组)。采用大鼠线栓法右侧中动脉栓塞模型(MCAO模型),于模型90 min时拔出线栓进行再灌注,再灌注同时,I/R组腹腔给予生理盐水10 m L/kg,HS组腹腔给予氢饱和生理盐水10 m L/kg。24 h后,对各组大鼠进行神经功能缺陷评分。断头取脑后,TTC染色法检测脑组织梗死体积(n=10)。选取缺血半暗带处大脑皮层组织进行相关指标测定(n=10)。HE染色观察大鼠缺血半暗带脑组织形态结构,测定缺血半暗带区域氧化应激反应,选取指标为SOD,MDA;并观察缺血半暗带区域炎症反应,利用Elisa方法测定该处TNF-α,IL-6含量。结果:TTC染色证实,右侧MCAO明显诱导大鼠脑局灶性缺血(P<0.05)。与I/R组相比,HS组明显降低脑梗死体积(P<0.05),显著降低大鼠神经功能缺陷评分(P<0.05)。与I/R组相比,氢饱和生理盐水治疗后,很好的保持了缺血半暗带区域细胞结构完整性,明显提高了SOD含量(P<0.05),有效降低了MDA水平(P<0.05),并且减轻了炎症因子TNF-α,IL-6含量(P<0.05)。结论:氢饱和生理盐水可有效的治疗脑缺血再灌注损伤,其机制可能涉及氢气在缺血半暗带区域的选择性的抗氧化应激作用,以及抗炎症反应。
Objective: The aim of this research was to study the effects on hydrogen-rich saline against the rat model of cerebral ischemia-reperfusion injury and elucidate the possible mechanisms. Methods: Sprague-Dawley rats weighing 250-300 g were distributed randomly into 3 groups(n=20): sham group,cerebral ischemia-reperfusion injury group(I/R group),and hydrogen-rich saline treated group(HS group). The model was induced by right middle cerebral artery occlusion(MCAO) via suture. Reperfusion was accomplished by withdrawing the suture after 90 min of ischemia. During the process of reperfusion,rats in I/R group were injected saline(10 m L/kg)intraperitoneally,and rats in HS group were injected hydrogen-rich saline(10 m L/kg) intraperitoneally. Neurological deficit scores were evaluated at 24 h after reperfusion. Then,the rats were decapitated,TTC tests were performed to detect the infarct volume of cortex(n=10). Ischemic penumbra was collected to analyze the involved indexes(n=10). We used HE staining to observe the morphology of cortical ischemic penumbra,and used SOD and MDA to evaluate the oxidative response in the region of cortical ischemic penumbra,and used TNF-α and IL-6 to evaluate the inflammatory response in cortical ischemic penumbra via Elisa test. Results: TTC staining test suggested that the model of right MCAO could induce focal ischemia of brain obviously(P<0.05). Compared with I/R rats,rats in HS group had decreased infarct volume of cortex(P<0.05) and lower neurological deficit scores(P<0.05). Rats in HS group maintained cellular structural integrity superiorly and had higher concentration of SOD(P<0.05),and lower concentration of MDA(P<0.05),TNF-α and IL-6(P<0.05) when compared to rats in I/R group. Conclusions: Hydrogen-rich saline could treat cerebral ischemia-reperfusion injury effectively,and the possible mechanisms involved the selectively antioxidant stress damage and effects of anti-inflammation in the region of cortical ischemic penumbra.
Objective: The aim of this research was to study the effects on hydrogen-rich saline against the rat model of cerebral ischemia-reperfusion injury and elucidate the possible mechanisms. Methods: Sprague-Dawley rats weighing 250-300 g were distributed randomly into 3 groups(n=20): sham group,cerebral ischemia-reperfusion injury group(I/R group),and hydrogen-rich saline treated group(HS group). The model was induced by right middle cerebral artery occlusion(MCAO) via suture. Reperfusion was accomplished by withdrawing the suture after 90 min of ischemia. During the process of reperfusion,rats in I/R group were injected saline(10 m L/kg)intraperitoneally,and rats in HS group were injected hydrogen-rich saline(10 m L/kg) intraperitoneally. Neurological deficit scores were evaluated at 24 h after reperfusion. Then,the rats were decapitated,TTC tests were performed to detect the infarct volume of cortex(n=10). Ischemic penumbra was collected to analyze the involved indexes(n=10). We used HE staining to observe the morphology of cortical ischemic penumbra,and used SOD and MDA to evaluate the oxidative response in the region of cortical ischemic penumbra,and used TNF-α and IL-6 to evaluate the inflammatory response in cortical ischemic penumbra via Elisa test. Results: TTC staining test suggested that the model of right MCAO could induce focal ischemia of brain obviously(P<0.05). Compared with I/R rats,rats in HS group had decreased infarct volume of cortex(P<0.05) and lower neurological deficit scores(P<0.05). Rats in HS group maintained cellular structural integrity superiorly and had higher concentration of SOD(P<0.05),and lower concentration of MDA(P<0.05),TNF-α and IL-6(P<0.05) when compared to rats in I/R group. Conclusions: Hydrogen-rich saline could treat cerebral ischemia-reperfusion injury effectively,and the possible mechanisms involved the selectively antioxidant stress damage and effects of anti-inflammation in the region of cortical ischemic penumbra.
引文
[1]IOhsawa,M Ishikawa,K Takahashi,et al.Hydrogen acts as a therapeutic antioxidant by selectively reducing cytotoxic oxygen radicals[J].Nat Med,2007,13(6):688-694
[2]Q Li,P Yu,Q Zeng,et al.Neuroprotective effect of hydrogen-rich saline in global cerebral ischemia/reperfusion rats:up-regulated tregs and down-regulated mir-21,mir-210 and nf-kappab expression[J].Neurochem Res,2016,2016:1-11
[3]YQ Liu,YF Liu,XM Ma,et al.Hydrogen-rich saline attenuates skin ischemia/reperfusion induced apoptosis via regulating bax/bcl-2 ratio and ask-1/jnk pathway[J].J Plast Reconstr Aesthet Surg,2015,68(7):147-156
[4]T Huang,W Wang,C Tu,et al.Hydrogen-rich saline attenuates ischemia-reperfusion injury in skeletal muscle[J].J Surg Res,2015,194(2):471-480
[5]DY Song,KM Oh,HN Yu,et al.Role of activating transcription factor3 in ischemic penumbra region following transient middle cerebral artery occlusion and reperfusion injury[J].Neurosci Res,2011,70(4):428-434
[6]刘漪,李雪梅,谭永星.氢气饱和生理盐水对局灶性脑缺血再灌注损伤大鼠神经功能缺损,神经细胞凋亡及凋亡相关蛋白表达的影响[J].临床神经病学杂志,2013,26(5):351-354Liu Yi,Li Xue-mei,Tan Yong-xing.Effects of saturated hydrogen saline on neurological deficit,neuronal apoptosis and the expression of apoptosis related proteins in rats with focal cerebral ischemia reperfusion injury[J].Journal of clinical neurology,2013,26(5):351-354
[7]X Zheng,Y Mao,J Cai,et al.Hydrogen-rich saline protects against intestinal ischemia/reperfusion injury in rats[J].Free Radic Res,2009,43(5):478-484
[8]EZ Longa,PR Weinstein,S Carlson,et al.Reversible middle cerebral artery occlusion without craniectomy in rats[J].Stroke,1989,20(1):84-91
[9]DC Rogers,CA Campbell,JL Stretton,et al.Correlation between motor impairment and infarct volume after permanent and transient middle cerebral artery occlusion in the rat[J].Stroke,1997,28(10):2060-2065
[10]SM Ostojic.Molecular hydrogen in sports medicine:New therapeutic perspectives[J].Int J Sports Med,2015,36(4):273-279
[11]J Li,Z Hong,H Liu,et al.Hydrogen-rich saline promotes the recovery of renal function after ischemia/reperfusion injury in rats via antiapoptosis and anti-inflammation[J].Front Pharmacol,2016,7:106
[12]ZL Luo,L Cheng,JD Ren.Hydrogen-rich saline protects against ischemia/reperfusion injury in grafts after pancreas transplantations by reducing oxidative stress in rats[J].Mediators Inflamm,2015,2015:1-8
[13]CB Zhang,YC Tang,XJ Xu,et al.Hydrogen gas inhalation protects against liver ischemia/reperfusion injury by activating the nf-kappab signaling pathway[J].Exp Ther Med,2015,9(6):2114-2120
[14]T Kawamura,N Wakabayashi,N Shigemura,et al.Hydrogen gas reduces hyperoxic lung injury via the nrf2 pathway in vivo[J].Am J Physiol Lung Cell Mol Physiol,2013,304(10):646-656
[15]Y Zhang,Q Sun,B He,et al.Anti-inflammatory effect of hydrogenrich saline in a rat model of regional myocardial ischemia and reperfusion[J].Int J Cardiol,2011,148(1):91-95
[16]Y Cui,H Zhang,M Ji,et al.Hydrogen-rich saline attenuates neuronal ischemia--reperfusion injury by protecting mitochondrial function in rats[J].J Surg Res,2014,192(2):564-572
[17]J Li,Y Dong,H Chen,et al.Protective effects of hydrogen-rich saline in a rat model of permanent focal cerebral ischemia via reducing oxidative stress and inflammatory cytokines[J].Brain Res,2012,1486:103-111
[18]BS Allen,GD Buckberg.Studies of isolated global brain ischaemia:I.Overview of irreversible brain injury and evolution of a new conceptredefining the time of brain death[J].Eur J Cardiothorac Surg,2012,41(5):1132-1137
[19]NJ Alkayed,I Harukuni,AS Kimes,et al.Gender-linked brain injury in experimental stroke[J].Stroke,1998,29(1):159-165
[20]WS Smith,GSung,S Starkman,et al.Safety and efficacy of mechanical embolectomy in acute isc hemic stroke:Results of the merci trial[J].Stroke,2005,36(7):1432-1438
[21]NJ Mac Dougall,KW Muir.Hyperglycaemia and infarct size in animal models of middle cerebral artery occlusion:Systematic review and meta-analysis[J].J Cereb Blood Flow Metab,2011,31(3):807-818
[22]AE Badr,W Yin,G Mychaskiw,et al.Dual effect of hbo on cerebral infarction in mcao rats[J].Am J Physiol Regul Integr Comp Physiol,2001,280(3):766-770
[23]Y Horita,O Honmou,K Harada,et al.Intravenous administration of glial cell line-derived neurotrophic factor gene-modified human mesenchymal stem cells protects against injury in a cerebral ischemia model in the adult rat[J].J Neurosci Res,2006,84(7):1495-1504
[24]N Shimamura,G Matchett,T Tsubokawa,et al.Comparison of siliconcoated nylon suture to plain nylon suture in the rat middle cerebral artery occlusion model[J].J Neurosci Methods,2006,156(1-2):161-165
[25]K Fukuda,S Asoh,M Ishikawa,et al.Inhalation of hydrogen gas suppresses hepatic injury caused by ischemia/reperfusion through reducing oxidative stress[J].Biochem Biophys Res Commun,2007,361(3):670-674
[26]Y Huang,K Xie,J Li,et al.Beneficial effects of hydrogen gas against spinal cord ischemia-reperfusion injury in rabbits[J].Brain Res,2011,1378:125-136
[27]BM Buchholz,DJ Kaczorowski,R Sugimoto,et al.Hydrogen inhalation ameliorates oxidative stress in transplantation induced intestinal graft injury[J].A Transplant,2008,8(10):2015-2024
[28]K Nagatani,K Wada,S Takeuchi,et al.Effect of hydrogen gas on the survival rate of mice following global cerebral ischemia[J].Shock,2012,37(6):645-652
[29]K Esposito,F Nappo,R Marfella,et al.Inflammatory cytokine concentrations are acutely increased by hyperglycemia in humans:Role of oxidative stress[J].Circulation,2002,106(16):2067-2072
[30]KA Kaminski,TA Bonda,JKorecki,et al.Oxidative stress and neutrophil activation-the two keystones of ischemia/reperfusion injury[J].Int J Cardiol,2002,86(1):41-59
[2]Q Li,P Yu,Q Zeng,et al.Neuroprotective effect of hydrogen-rich saline in global cerebral ischemia/reperfusion rats:up-regulated tregs and down-regulated mir-21,mir-210 and nf-kappab expression[J].Neurochem Res,2016,2016:1-11
[3]YQ Liu,YF Liu,XM Ma,et al.Hydrogen-rich saline attenuates skin ischemia/reperfusion induced apoptosis via regulating bax/bcl-2 ratio and ask-1/jnk pathway[J].J Plast Reconstr Aesthet Surg,2015,68(7):147-156
[4]T Huang,W Wang,C Tu,et al.Hydrogen-rich saline attenuates ischemia-reperfusion injury in skeletal muscle[J].J Surg Res,2015,194(2):471-480
[5]DY Song,KM Oh,HN Yu,et al.Role of activating transcription factor3 in ischemic penumbra region following transient middle cerebral artery occlusion and reperfusion injury[J].Neurosci Res,2011,70(4):428-434
[6]刘漪,李雪梅,谭永星.氢气饱和生理盐水对局灶性脑缺血再灌注损伤大鼠神经功能缺损,神经细胞凋亡及凋亡相关蛋白表达的影响[J].临床神经病学杂志,2013,26(5):351-354Liu Yi,Li Xue-mei,Tan Yong-xing.Effects of saturated hydrogen saline on neurological deficit,neuronal apoptosis and the expression of apoptosis related proteins in rats with focal cerebral ischemia reperfusion injury[J].Journal of clinical neurology,2013,26(5):351-354
[7]X Zheng,Y Mao,J Cai,et al.Hydrogen-rich saline protects against intestinal ischemia/reperfusion injury in rats[J].Free Radic Res,2009,43(5):478-484
[8]EZ Longa,PR Weinstein,S Carlson,et al.Reversible middle cerebral artery occlusion without craniectomy in rats[J].Stroke,1989,20(1):84-91
[9]DC Rogers,CA Campbell,JL Stretton,et al.Correlation between motor impairment and infarct volume after permanent and transient middle cerebral artery occlusion in the rat[J].Stroke,1997,28(10):2060-2065
[10]SM Ostojic.Molecular hydrogen in sports medicine:New therapeutic perspectives[J].Int J Sports Med,2015,36(4):273-279
[11]J Li,Z Hong,H Liu,et al.Hydrogen-rich saline promotes the recovery of renal function after ischemia/reperfusion injury in rats via antiapoptosis and anti-inflammation[J].Front Pharmacol,2016,7:106
[12]ZL Luo,L Cheng,JD Ren.Hydrogen-rich saline protects against ischemia/reperfusion injury in grafts after pancreas transplantations by reducing oxidative stress in rats[J].Mediators Inflamm,2015,2015:1-8
[13]CB Zhang,YC Tang,XJ Xu,et al.Hydrogen gas inhalation protects against liver ischemia/reperfusion injury by activating the nf-kappab signaling pathway[J].Exp Ther Med,2015,9(6):2114-2120
[14]T Kawamura,N Wakabayashi,N Shigemura,et al.Hydrogen gas reduces hyperoxic lung injury via the nrf2 pathway in vivo[J].Am J Physiol Lung Cell Mol Physiol,2013,304(10):646-656
[15]Y Zhang,Q Sun,B He,et al.Anti-inflammatory effect of hydrogenrich saline in a rat model of regional myocardial ischemia and reperfusion[J].Int J Cardiol,2011,148(1):91-95
[16]Y Cui,H Zhang,M Ji,et al.Hydrogen-rich saline attenuates neuronal ischemia--reperfusion injury by protecting mitochondrial function in rats[J].J Surg Res,2014,192(2):564-572
[17]J Li,Y Dong,H Chen,et al.Protective effects of hydrogen-rich saline in a rat model of permanent focal cerebral ischemia via reducing oxidative stress and inflammatory cytokines[J].Brain Res,2012,1486:103-111
[18]BS Allen,GD Buckberg.Studies of isolated global brain ischaemia:I.Overview of irreversible brain injury and evolution of a new conceptredefining the time of brain death[J].Eur J Cardiothorac Surg,2012,41(5):1132-1137
[19]NJ Alkayed,I Harukuni,AS Kimes,et al.Gender-linked brain injury in experimental stroke[J].Stroke,1998,29(1):159-165
[20]WS Smith,GSung,S Starkman,et al.Safety and efficacy of mechanical embolectomy in acute isc hemic stroke:Results of the merci trial[J].Stroke,2005,36(7):1432-1438
[21]NJ Mac Dougall,KW Muir.Hyperglycaemia and infarct size in animal models of middle cerebral artery occlusion:Systematic review and meta-analysis[J].J Cereb Blood Flow Metab,2011,31(3):807-818
[22]AE Badr,W Yin,G Mychaskiw,et al.Dual effect of hbo on cerebral infarction in mcao rats[J].Am J Physiol Regul Integr Comp Physiol,2001,280(3):766-770
[23]Y Horita,O Honmou,K Harada,et al.Intravenous administration of glial cell line-derived neurotrophic factor gene-modified human mesenchymal stem cells protects against injury in a cerebral ischemia model in the adult rat[J].J Neurosci Res,2006,84(7):1495-1504
[24]N Shimamura,G Matchett,T Tsubokawa,et al.Comparison of siliconcoated nylon suture to plain nylon suture in the rat middle cerebral artery occlusion model[J].J Neurosci Methods,2006,156(1-2):161-165
[25]K Fukuda,S Asoh,M Ishikawa,et al.Inhalation of hydrogen gas suppresses hepatic injury caused by ischemia/reperfusion through reducing oxidative stress[J].Biochem Biophys Res Commun,2007,361(3):670-674
[26]Y Huang,K Xie,J Li,et al.Beneficial effects of hydrogen gas against spinal cord ischemia-reperfusion injury in rabbits[J].Brain Res,2011,1378:125-136
[27]BM Buchholz,DJ Kaczorowski,R Sugimoto,et al.Hydrogen inhalation ameliorates oxidative stress in transplantation induced intestinal graft injury[J].A Transplant,2008,8(10):2015-2024
[28]K Nagatani,K Wada,S Takeuchi,et al.Effect of hydrogen gas on the survival rate of mice following global cerebral ischemia[J].Shock,2012,37(6):645-652
[29]K Esposito,F Nappo,R Marfella,et al.Inflammatory cytokine concentrations are acutely increased by hyperglycemia in humans:Role of oxidative stress[J].Circulation,2002,106(16):2067-2072
[30]KA Kaminski,TA Bonda,JKorecki,et al.Oxidative stress and neutrophil activation-the two keystones of ischemia/reperfusion injury[J].Int J Cardiol,2002,86(1):41-59