促红细胞生成素对大鼠肝缺血再灌注损伤的保护作用及基质金属蛋白酶-9和半胱天冬酶-1表达的影响
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摘要
目的:探讨促红细胞生成素(EPO)对大鼠肝缺血再灌注损伤后的保护作用及对肝基质金属蛋白酶-9(MMP-9)和半胱天冬酶-1(caspase-1)表达的影响。方法:将健康雄性SD大鼠随机分为5组,缺血再灌注损伤组(I/R)、促红细胞生成素低剂量组(L组)、中剂量组(M组)、高剂量组(H组)和假手术组。H-E染色观察各组肝组织的病理变化;免疫组织化学显色和免疫印迹法检测MMP-9、caspase-1蛋白表达;比色法检测血清中丙氨酸氨基转移酶(ALT)和天门冬氨酸氨基转移酶(AST)水平。结果:光镜下I/R组肝细胞出现水肿及片状坏死,大量炎性细胞浸润;EPO预处理组出现肝细胞水肿、坏死及炎细胞浸润程度明显减轻。与假手术组比较,缺血再灌注各组肝组织MMP-9和caspase-1表达水平均上升,血清ALT和AST水平也均上升,且均以I/R组上升最为明显。EPO不同剂量组间比较,中剂量组大鼠肝MMP-9和caspase-1表达水平、ALT和AST的水平与低、高剂量组比较均降低。结论:EPO对大鼠肝缺血再灌注损伤具有保护作用,最佳浓度是3000U/kg。该保护作用可能与抑制MMP-9、caspase-1介导的过度炎症反应有关。
Objective:To study the effect of erythropoietin(EPO)on hepatic ischemia reperfusion injury and the expression of hepatic matrix metalloproteinase 9(MMP-9)and cysteinyl aspartate specific proteinase-1(caspase-1).Methods:Fifty healthy male SD rats were randomly divided into 5 random groups:ischemia reperfusion group(I/R group),low-dose EPO group(L group),middle-dose EPO group(M group),high-dose EPO group(H group)and sham-operation group(sham group).The morphologic changes in the liver were observed by light microscope after H-E staining.The protein levels of MMP-9 and caspase-1 in the liver tissues were detected by immunohistochemical staining and Western blotting.Serum alanine transaminase(ALT)and aspartate aminotransferase(AST)levels were measured by methods recommended by the manufacturer.Results:Under light microscope,there were edema and necrosis of hepatocytes and a large number of inflammatory cell infiltration in I/R group.Meanwhile,necrosis and infiltration of inflammatory cells in EPO preconditioning group were significantly alleviated.Compared with the control group,the expression value of MMP-9 and caspase-1 in the liver and the activity of ALT and AST in serum of the other four groups increased(P<0.01),and the I/R group showed the most significant change.Compared with L and H groups,the expression of MMP-9 and caspase-1 in the liver of M group was lower than that of L and H groups.Conclusion:EPO has the protective effect on hepatic ischemia-reperfusion injury in rats and the best concentration is 3000 U/kg.The protective mechanism of EPO may be related to the inhibition of MMP-9 and caspase-1 which mediates over-inflammation.
Objective:To study the effect of erythropoietin(EPO)on hepatic ischemia reperfusion injury and the expression of hepatic matrix metalloproteinase 9(MMP-9)and cysteinyl aspartate specific proteinase-1(caspase-1).Methods:Fifty healthy male SD rats were randomly divided into 5 random groups:ischemia reperfusion group(I/R group),low-dose EPO group(L group),middle-dose EPO group(M group),high-dose EPO group(H group)and sham-operation group(sham group).The morphologic changes in the liver were observed by light microscope after H-E staining.The protein levels of MMP-9 and caspase-1 in the liver tissues were detected by immunohistochemical staining and Western blotting.Serum alanine transaminase(ALT)and aspartate aminotransferase(AST)levels were measured by methods recommended by the manufacturer.Results:Under light microscope,there were edema and necrosis of hepatocytes and a large number of inflammatory cell infiltration in I/R group.Meanwhile,necrosis and infiltration of inflammatory cells in EPO preconditioning group were significantly alleviated.Compared with the control group,the expression value of MMP-9 and caspase-1 in the liver and the activity of ALT and AST in serum of the other four groups increased(P<0.01),and the I/R group showed the most significant change.Compared with L and H groups,the expression of MMP-9 and caspase-1 in the liver of M group was lower than that of L and H groups.Conclusion:EPO has the protective effect on hepatic ischemia-reperfusion injury in rats and the best concentration is 3000 U/kg.The protective mechanism of EPO may be related to the inhibition of MMP-9 and caspase-1 which mediates over-inflammation.
引文
[1] Kapan M,Gumus M,Onder A,et al.The effects of ellagic acid on the liver and remote organs′oxidative stress and structure after hepatic ischemia reperfusion injury caused by pringle maneuver in rats[J].Bratisl Lek Listy,2012,113(5):274-281.
[2] Li J,Li R J,Lv G Y,et al.The mechanisms and strategies to protect from hepatic ischemia reperfusion injury[J].Eur Rev Med Pharmacol Sci,2015,19(11):2036-2047.
[3] Guan L Y.Mechanisms of hepatic ischemia-reperfusion injury and protective effects of nitric oxide[J].World J Gastrointest Surg,2014,6(7):122-128
[4] Philippe V L,Claude L.Chemokine and cytokine processing by matrix metalloproteinases and its effect on leukocyte migration and inflammation[J].J Leukoc Biol,2007,82(6):1375-1381.
[5] Parks W C, Wilson C L, López-boado Y S. Matrix metalloproteinases as modulators of inflammation and innate immunity[J].Nat Rev Immunol,2004,4(8):617-629.
[6] Maximilian S,Gerhard H,Veravoorn A.Neumann erythropoietin reduces ischemia-reperfusion injury after liver transplantation in rats[J].Eur Soc Organ Transplantat,2009,22(5):738-746.
[7] Gollg Y, Xue B,Jiao J,et al.Triptolideinhibits COX-2expression and PGE2release by suppressing the activity of NFkappaB and JNK in LPS-treated microglia[J].J Neurochem,2008,107(3):779-788.
[8] Li J,Yuan T,Zhao X,et al.Protective effects of sevoflurane in hepatic ischemia-reperfusion injury[J].Int J Immunopathol Pharmacol,2016,29(2):300-307.
[9] Ferchichi H,Bacha S,Kourda N,et al.Animal model of liver ischemia reperfusion:biochemical and histological evaluation[J].Tunis Me,2016,94(3):235-243.
[10]张志斌,朱志军.炎症反应在肝脏缺血再灌注损伤中的作用研究进展[J].山东医药,2014,54(15):101-103.
[11] Andrej K,Kessler J S,Marc H,et al.Matrix metalloproteinase-9promotes neutrophil and T cell recruitment and migration in the postischemic liver[J].J Leukoc Biol,2006,79(6):1295-1305.
[12] TakashiH, ConstantinoF, BusuttilRW, etal.Metalloproteinase-9deficiency protects against hepatic ischemia/reperfusion injury[J].Hepatology,2008,47(1):186-198.
[13] Coito AJ.Leukocyte transmigration across endothelial and extracellular matrix protein barriers in liver ischemia/reperfusion injury[J].Curr Opin Organ Transplant,2011,16(1):34-40.
[14] De Nardo D,Latz E.NLRP3inflammasomes link inflammation and metabolic disease[J].Trends Immunol,2011,32(8):373-379.
[15] Schroder K,Tschopp J.The inflammasomes.Cell,2010,140(6):821-832.
[16] Brines M, Cerami A. Discovering erythropoietin′s extrahematopoietic functions:biology and clinical promise[J].Kidney Int,2006,70(2):246-250.
[17] Joyeux-Faure M.Cellular protection by erythropoietin:new therapeutic implications?[J].J Pharmacol Exp Ther,2007,323(3):759-762.
[18] Mocini D,Leone T,Tubaro M,et al.Structure,production and function of erythropoietin:implications for therapeutical use in cardiovascular disease[J].Curr Med Chem,2007,14(21):2278-2287
[19]陈茂松,王军,欧雷.促红细胞生成素对大鼠肝脏缺血再灌注损伤的抗炎作用[J].广东医学,2016,37(6):828-830.
[20] Li R,Zhang L M,Sun W B.Erythropoietin rescues primary rat cortical neurons from pyroptosis and apoptosis via Erk1/2-Nrf2/Bach1signal pathway[J].Brain Res Bull,2017,130:236-244.
[2] Li J,Li R J,Lv G Y,et al.The mechanisms and strategies to protect from hepatic ischemia reperfusion injury[J].Eur Rev Med Pharmacol Sci,2015,19(11):2036-2047.
[3] Guan L Y.Mechanisms of hepatic ischemia-reperfusion injury and protective effects of nitric oxide[J].World J Gastrointest Surg,2014,6(7):122-128
[4] Philippe V L,Claude L.Chemokine and cytokine processing by matrix metalloproteinases and its effect on leukocyte migration and inflammation[J].J Leukoc Biol,2007,82(6):1375-1381.
[5] Parks W C, Wilson C L, López-boado Y S. Matrix metalloproteinases as modulators of inflammation and innate immunity[J].Nat Rev Immunol,2004,4(8):617-629.
[6] Maximilian S,Gerhard H,Veravoorn A.Neumann erythropoietin reduces ischemia-reperfusion injury after liver transplantation in rats[J].Eur Soc Organ Transplantat,2009,22(5):738-746.
[7] Gollg Y, Xue B,Jiao J,et al.Triptolideinhibits COX-2expression and PGE2release by suppressing the activity of NFkappaB and JNK in LPS-treated microglia[J].J Neurochem,2008,107(3):779-788.
[8] Li J,Yuan T,Zhao X,et al.Protective effects of sevoflurane in hepatic ischemia-reperfusion injury[J].Int J Immunopathol Pharmacol,2016,29(2):300-307.
[9] Ferchichi H,Bacha S,Kourda N,et al.Animal model of liver ischemia reperfusion:biochemical and histological evaluation[J].Tunis Me,2016,94(3):235-243.
[10]张志斌,朱志军.炎症反应在肝脏缺血再灌注损伤中的作用研究进展[J].山东医药,2014,54(15):101-103.
[11] Andrej K,Kessler J S,Marc H,et al.Matrix metalloproteinase-9promotes neutrophil and T cell recruitment and migration in the postischemic liver[J].J Leukoc Biol,2006,79(6):1295-1305.
[12] TakashiH, ConstantinoF, BusuttilRW, etal.Metalloproteinase-9deficiency protects against hepatic ischemia/reperfusion injury[J].Hepatology,2008,47(1):186-198.
[13] Coito AJ.Leukocyte transmigration across endothelial and extracellular matrix protein barriers in liver ischemia/reperfusion injury[J].Curr Opin Organ Transplant,2011,16(1):34-40.
[14] De Nardo D,Latz E.NLRP3inflammasomes link inflammation and metabolic disease[J].Trends Immunol,2011,32(8):373-379.
[15] Schroder K,Tschopp J.The inflammasomes.Cell,2010,140(6):821-832.
[16] Brines M, Cerami A. Discovering erythropoietin′s extrahematopoietic functions:biology and clinical promise[J].Kidney Int,2006,70(2):246-250.
[17] Joyeux-Faure M.Cellular protection by erythropoietin:new therapeutic implications?[J].J Pharmacol Exp Ther,2007,323(3):759-762.
[18] Mocini D,Leone T,Tubaro M,et al.Structure,production and function of erythropoietin:implications for therapeutical use in cardiovascular disease[J].Curr Med Chem,2007,14(21):2278-2287
[19]陈茂松,王军,欧雷.促红细胞生成素对大鼠肝脏缺血再灌注损伤的抗炎作用[J].广东医学,2016,37(6):828-830.
[20] Li R,Zhang L M,Sun W B.Erythropoietin rescues primary rat cortical neurons from pyroptosis and apoptosis via Erk1/2-Nrf2/Bach1signal pathway[J].Brain Res Bull,2017,130:236-244.