马栗种子提取物通过抑制活性氧及JNK途径保护刀豆蛋白A诱导的小鼠急性肝损伤
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摘要
目的观察马栗种子提取物(AH)对刀豆蛋白A(Con A)诱导的小鼠急性肝损伤的保护作用,并探讨其相关机制是否与AH抑制氧化应激及其介导的c-Jun氨基末端激酶(JNK)途径有关。方法小鼠尾静脉一次性注射Con A(20mg·kg~(-1))制备急性肝损伤模型,AH保护组分别给予12.5、25、50 mg·kg~(-1)AH连续灌胃20 d预保护;全自动生化分析仪检测血清ALT、AST、TP、Alb、A/G;ELISA法检测小鼠血清IFN-γ、TNF-α水平;肝组织HE染色并进行病理学损伤分级评估;试剂盒检测肝组织MDA、SOD、GSH水平;分光光度法检测caspase-3活性;Western blot检测caspase-3、Bax、Bcl-2、p-JNK、p-Akt的蛋白表达情况。结果与Con A模型组相比,不同浓度的AH处理组小鼠血清ALT、AST、IFN-γ、TNF-α水平明显降低,TP、Alb、A/G比明显增高;肝组织的MDA水平明显降低,SOD、GSH水平明显升高。肝组织病理学与血清学指标改变相一致,AH处理组相比Con A模型组病理损伤明显减轻,病理损伤分级降低;与模型组相比,AH保护组细胞色素C、caspase-3、Bax/Bcl-2、p-JNK均明显降低,而pAkt蛋白表达升高。结论 AH可明显改善Con A诱导的小鼠急性肝损伤,其机制与AH抗氧化应激,进而抑制JNK/线粒体凋亡途径有关。
Aim To investigate the effect of Aesculus hippocastanum seed extract( AH) on concanavalin A( Con A)-induced acute liver injury in mice,and to explore whether the mechanism was related to the inhibitory effect of AH on oxidative stress and c-Jun N-terminal kinase( JNK). Methods Con A( 20 mg ·kg~(-1))was administered via tail vein injecting to induce hepatic damage in mice. The groups of AH were given at12. 5,25,50 mg·kg~(-1) by oral gavage separately for20 days. The serum levels of AST,ALT,TP,and Alb were determined by automatic biochemical analyzer and the A/G ratio was calculated. TNF-α and IFN-γ levels were assayed by ELISA. The liver tissue was attained by HE and the histopathological changes were calculated. The MDA,SOD,GSH contents of liver tissues were assayed by related kits. The activity of caspase-3 was detected by spectrophotometry. The expressions of cytochrome C and Bax,Bcl-2,p-JNK and p-Akt were detected by Western blot. Results The serum levels of ALT,AST,IFN-γ and TNF-α in AH groups were significantly lower than those in Con A-injured group,while the levels of TP,Alb and A/G were significantly higher. The SOD and GSH levels of liver tissues significantly increased and MDA level decreased; liver histopathological changes were consistent with those of the serological indicators,and AH treatment significantly reduced the pathological damage induced by Con A. In AH group,the expression of cytochrome C,caspase-3,Bax/Bcl-2 ratio and p-JNK markedly decreased,while the expression of p-Akt protein increased compared with Con A model group. Conclusion AH could significantly protect the Con A-induced acute liver injury in mice via inhibition of ROS and JNK pathway.
Aim To investigate the effect of Aesculus hippocastanum seed extract( AH) on concanavalin A( Con A)-induced acute liver injury in mice,and to explore whether the mechanism was related to the inhibitory effect of AH on oxidative stress and c-Jun N-terminal kinase( JNK). Methods Con A( 20 mg ·kg~(-1))was administered via tail vein injecting to induce hepatic damage in mice. The groups of AH were given at12. 5,25,50 mg·kg~(-1) by oral gavage separately for20 days. The serum levels of AST,ALT,TP,and Alb were determined by automatic biochemical analyzer and the A/G ratio was calculated. TNF-α and IFN-γ levels were assayed by ELISA. The liver tissue was attained by HE and the histopathological changes were calculated. The MDA,SOD,GSH contents of liver tissues were assayed by related kits. The activity of caspase-3 was detected by spectrophotometry. The expressions of cytochrome C and Bax,Bcl-2,p-JNK and p-Akt were detected by Western blot. Results The serum levels of ALT,AST,IFN-γ and TNF-α in AH groups were significantly lower than those in Con A-injured group,while the levels of TP,Alb and A/G were significantly higher. The SOD and GSH levels of liver tissues significantly increased and MDA level decreased; liver histopathological changes were consistent with those of the serological indicators,and AH treatment significantly reduced the pathological damage induced by Con A. In AH group,the expression of cytochrome C,caspase-3,Bax/Bcl-2 ratio and p-JNK markedly decreased,while the expression of p-Akt protein increased compared with Con A model group. Conclusion AH could significantly protect the Con A-induced acute liver injury in mice via inhibition of ROS and JNK pathway.
引文
[1]刘慧敏,韩延忠,郭玉明,等.六味五灵片对刀豆蛋白A诱导的小鼠急性免疫性肝损伤的保护作用研究[J].中国药理学通报,2017,33(1):133-40.[1]Liu H M,Han Y Z,Guo Y M,et al.The protection effects of Liuweiwuling Tablets against concanavalin A-induced acute immunological liver injury in mice[J].Chin Pharmacol Bull,2017,33(1):133-40.
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[8]Selvakumar G P,Manivasagam T,Rekha K R,et al.Escin,a novel triterpene,mitigates chronic MPTP/p-induced dopaminergic toxicity by attenuating mitochondrial dysfunction,oxidative stress,and apoptosis[J].J Mol Neurosci,2015,55(1):184-97.
[9]Masaki H,Sakaki S,Atsumi T,et al.Active-oxygen scavenging activity of plant extracts[J].Biol Pharm Bull,1995,18(1):162-6.
[10]VakováJ,FejerˇcákováA,MojˇziˇsováG,et al.Antioxidant potential of Aesculus hippocastanum extract and escin against reactive oxygen and nitrogen species[J].Eur Rev Med Pharmacol Sci,2015,19(5):879-86.
[11]Hu X M,Zhang Y,Zeng F D.Effects of beta-aescin on apoptosis induced by transient focal cerebral ischemia in rats[J].Acta Pharmacol Sin,2004,25(10):1267-75.
[12]Elmas O,Erbas O,Yigitturk G.The efficacy of Aesculus hippocastanum seeds on diabetic nephropathy in a streptozotocin-induced diabetic rat model[J].Biomed Pharmacother,2016,83:392-6.
[13]杨社华,葛斌,王艳红,等.马栗籽提取物对急性肝损伤小鼠自由基及TNF-α含量的影响[J].中国医院药学杂志,2014,34(17):1453-6.[13]Yang S H,Ge B,Wang Y H,et al.Extracts of European Hippocastanum seeds carbon tetrachloride liver injury free radicals TNF-α[J].Chin J Hosp Pharm,2014,34(17):1453-6.
[14]Wang F,Xue Y,Yang J,et al.Hepatoprotective effect of apple polyphenols against concanavalin A-induced immunological liver injury in mice[J].Chem Biol Interact,2016,258:159-65.
[15]Horwacik I,Rokita H.Targeting of tumor-associated gangliosides with antibodies affects signaling pathways and leads to cell death including apoptosis[J].Apoptosis,2015,20(5):679-88.
[16]Linton M F,Babaev V R,Huang J,et al.Macrophage apoptosis and efferocytosis in the pathogenesis of atherosclerosis[J].Circ J,2016,80(11):2259-68.
[2]杨扬,吴小琴,李小枫,等.栀子柏皮汤及含栀子配伍组对免疫性肝损伤小鼠的保护作用[J].中国药理学通报,2015,31(12):1764-9.[2]Yang Y,Wu X Q,Li X F,et al.Study on the protective effect in immunological liver injury in mice of Zhizi Baipi Soup and its disassembled prescription[J].Chin Pharmacol Bull,2015,31(12):1764-9.
[3]Schwabe R F,Brenner D A.Mechanisms of liver injury.I.TNFalpha-induced liver injury:role of IKK,JNK,and ROS pathways[J].Am J Physiol Gastrointest Liver Physiol,2006,290(4):G583-9.
[4]Maeda S,Chang L,Li Z W,et al.IKKbeta is required for prevention of apoptosis mediated by cell-bound but not by circulating TNFalpha[J].Immunity,2003,19(5):725-37.
[5]Streetz K,Fregien B,Plumpe J,et al.Dissection of the intracellular pathways in hepatocytes suggests a role for Jun kinase and IFN regulatory factor-1 in Con A-induced liver failure[J].J Immunol,2001,167(1):514-23.
[6]Sirtori C R.Aescin:pharmacology,pharmacokinetics and therapeutic profile[J].Pharmacol Res,2001,44(3):183-93.
[7]Kü9ükkurt I,Ince S,Kele爧H,et al.Beneficial effects of Aesculus hippocastanum L.seed extract on the body’s own antioxidant defense system on subacute administration[J].J Ethnopharmacol,2010,129(1):18-22.
[8]Selvakumar G P,Manivasagam T,Rekha K R,et al.Escin,a novel triterpene,mitigates chronic MPTP/p-induced dopaminergic toxicity by attenuating mitochondrial dysfunction,oxidative stress,and apoptosis[J].J Mol Neurosci,2015,55(1):184-97.
[9]Masaki H,Sakaki S,Atsumi T,et al.Active-oxygen scavenging activity of plant extracts[J].Biol Pharm Bull,1995,18(1):162-6.
[10]VakováJ,FejerˇcákováA,MojˇziˇsováG,et al.Antioxidant potential of Aesculus hippocastanum extract and escin against reactive oxygen and nitrogen species[J].Eur Rev Med Pharmacol Sci,2015,19(5):879-86.
[11]Hu X M,Zhang Y,Zeng F D.Effects of beta-aescin on apoptosis induced by transient focal cerebral ischemia in rats[J].Acta Pharmacol Sin,2004,25(10):1267-75.
[12]Elmas O,Erbas O,Yigitturk G.The efficacy of Aesculus hippocastanum seeds on diabetic nephropathy in a streptozotocin-induced diabetic rat model[J].Biomed Pharmacother,2016,83:392-6.
[13]杨社华,葛斌,王艳红,等.马栗籽提取物对急性肝损伤小鼠自由基及TNF-α含量的影响[J].中国医院药学杂志,2014,34(17):1453-6.[13]Yang S H,Ge B,Wang Y H,et al.Extracts of European Hippocastanum seeds carbon tetrachloride liver injury free radicals TNF-α[J].Chin J Hosp Pharm,2014,34(17):1453-6.
[14]Wang F,Xue Y,Yang J,et al.Hepatoprotective effect of apple polyphenols against concanavalin A-induced immunological liver injury in mice[J].Chem Biol Interact,2016,258:159-65.
[15]Horwacik I,Rokita H.Targeting of tumor-associated gangliosides with antibodies affects signaling pathways and leads to cell death including apoptosis[J].Apoptosis,2015,20(5):679-88.
[16]Linton M F,Babaev V R,Huang J,et al.Macrophage apoptosis and efferocytosis in the pathogenesis of atherosclerosis[J].Circ J,2016,80(11):2259-68.
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