鸡血藤总黄酮对大肠杆菌败血症的治疗作用
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摘要
为研究鸡血藤总黄酮对小鼠大肠杆菌败血症的治疗作用,本研究采用腹腔注射大肠杆菌菌液的方法建立小鼠大肠杆菌败血症模型,1 h后灌胃给予不同剂量鸡血藤总黄酮,12 h后观察小鼠临床表现并进行评分,然后剖杀小鼠,比较肝脏、胸腺和脾脏的外观、脏器指数和肝脏病理切片结果,检测各项血液常规指标,分析血清中谷草转氨酶(AST)、谷丙转氨酶(ALT)及乳酸脱氢酶(LDH)水平,测定血清中白细胞介素1β(IL-1β)、IL-2、IL-6、IL-10及肿瘤坏死因子α(TNF-α)等炎性细胞因子的水平。结果发现,腹腔注射大肠杆菌后引起小鼠被毛粗乱、精神沉郁、眼睛红肿、眼部分泌物增加且呼吸急促等症状,部分严重的小鼠出现抽搐、休克现象,肝脏、胸腺、脾脏出现萎缩、变小的现象,且脏器指数下降。肝脏病理切片可观察到肝索紊乱、肝窦增大、细胞边缘不清晰、部分区域炎性细胞浸润。血液中白细胞(WBC)、淋巴细胞(LYM)、血红蛋白(HGB)、红细胞(RBC)和血小板(PLT)等指标水平发生显著改变(P<0.05),且血清中ALT、AST、LDH及炎性细胞因子等水平显著升高(P<0.05)。而鸡血藤总黄酮和阿米卡星处理可改善上述临床表现,抑制相关指标的改变,使其趋于与对照组相似水平。以上结果表明,鸡血藤总黄酮可通过提高机体免疫水平及抑制炎性反应减轻大肠杆菌败血症引起的机体损伤。
In order to evaluate the therapeutic effect of total flavonoids from Spatholobus suberectus Dunn(TFSD) on Escherichia coli(E.coli) sepsis,sepsis animal models were established in mice by intraperitoneally injected with E.coli,mice were administrated with various dose of TFSD after 1 h,the clinical manifestations of mice were observed and scored after 12 h.Then the mice were dissected,the liver,thymus and spleen were observed and weighted for the calculation of their organ indexes,pathological sections of livers were prepared for detail analysis of pathological change.Blood routine indicators were measured to analyze the levels of aspartate aminotransferase(AST),alanine aminotransferase(ALT) and lactate dehydrogenase(LDH) in serum.The levels of inflammatory cytokines such as interleukin-1β(IL-1β),IL-2,IL-6,IL-10 and tumor necrosis factor α(TNF-α) in serum were determined.The results showed that the injection of E.coli resulted in irregular hair coat,poor spirit,inflamed eyes,increase of secretion from eyes,tachypnea,even convulsion or shock.Atrophy was observed from liver,thymus and spleen tissues,corresponding with reduced organ index.Pathological sections indicated disorder of hepatic cord,swell of hepatic sinus,fuzziness of cell edges,and infiltration of inflammatory cells.The concentrations of leukocytes(WBC),lymphocyte(LYM),hemoglobin(HGB),red blood cell(RBC) and platelet(PLT) were significantly altered by the injection of E.coli(P<0.05).The levels of ALT,AST,LDH and inflammatory cytokines were significantly increased(P<0.05).However,the treatment of TFSD and amikacin was able to alleviate the clinical manifestation and inhibit the change of organ indexes,which tended to be similar to the control group.The results indicated that TFSD was able to alleviate E.coli induced injury by improving the immune response and inhibit inflammatory reaction in mice.
In order to evaluate the therapeutic effect of total flavonoids from Spatholobus suberectus Dunn(TFSD) on Escherichia coli(E.coli) sepsis,sepsis animal models were established in mice by intraperitoneally injected with E.coli,mice were administrated with various dose of TFSD after 1 h,the clinical manifestations of mice were observed and scored after 12 h.Then the mice were dissected,the liver,thymus and spleen were observed and weighted for the calculation of their organ indexes,pathological sections of livers were prepared for detail analysis of pathological change.Blood routine indicators were measured to analyze the levels of aspartate aminotransferase(AST),alanine aminotransferase(ALT) and lactate dehydrogenase(LDH) in serum.The levels of inflammatory cytokines such as interleukin-1β(IL-1β),IL-2,IL-6,IL-10 and tumor necrosis factor α(TNF-α) in serum were determined.The results showed that the injection of E.coli resulted in irregular hair coat,poor spirit,inflamed eyes,increase of secretion from eyes,tachypnea,even convulsion or shock.Atrophy was observed from liver,thymus and spleen tissues,corresponding with reduced organ index.Pathological sections indicated disorder of hepatic cord,swell of hepatic sinus,fuzziness of cell edges,and infiltration of inflammatory cells.The concentrations of leukocytes(WBC),lymphocyte(LYM),hemoglobin(HGB),red blood cell(RBC) and platelet(PLT) were significantly altered by the injection of E.coli(P<0.05).The levels of ALT,AST,LDH and inflammatory cytokines were significantly increased(P<0.05).However,the treatment of TFSD and amikacin was able to alleviate the clinical manifestation and inhibit the change of organ indexes,which tended to be similar to the control group.The results indicated that TFSD was able to alleviate E.coli induced injury by improving the immune response and inhibit inflammatory reaction in mice.
引文
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[12] 陈海兰,蒙西南,赵尉丹,等.鸡血藤总黄酮乙酸乙酯部位对大肠杆菌脂多糖(LPS)刺激RAW264.7免疫细胞氧化应激的调节作用[J].江苏农业科学,2017,45(8):154-156.CHEN H L,MENG X N,ZHAO W D,et al.Regulatory effect of ethyl acetate extract of total flavonoids of Caulis spatholobi on lipopolysaccharide induced oxidative stress in RAW264.7 cells[J].Jiangsu Agricultural Sciences,2017,45(8):154-156.(in Chinese)
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[14] 付远妨,赵尉丹,蒙西南,等.鸡血藤黄酮乙酸乙酯部位对H2O2诱导RAW264.7细胞氧化应激的调节作用[J].黑龙江畜牧兽医,2017,12:145-147.FU Y F,ZHAO W D,MENG X N,et al.Regulatory effect of ethyl acetate extract of total flavonoid of Spatholobus suberectus Dunn on the oxidative stress in H2O2 stimulated RAW264.7 cells[J].Heilongjiang Animal Science and Veterinary Medicine,2017,12:145-147.(in Chinese)
[15] 韩玉杰.白介素17(IL-17)在大肠埃希菌诱导的新生大鼠败血症中的意义[D].苏州:苏州大学,2012.HAN Y J.The significance of interleukin 17 (IL-17) in neonatal rats septicemia with E.coli[D].Suzhou:Suzhou University,2012.(in Chinese)
[16] 厉世伟,谷静娟,赵璐.金银花、连翘对大肠杆菌感染模型小鼠血清Th1/Th2细胞因子的调节作用[J].现代中西医结合杂志,2013,22(6):594-596.LI S W,GU J J,ZHAO L.Regulatory mechanism of Flos lonicerae and Fructus forsythia for mouse infected by Escherichia coli[J].Modern Journal of Integrated Chinese Traditional and Western Medicine,2013,22(6):594-596.(in Chinese)
[17] 刘灿,王婵,左之才,等.黄芪多糖对腹腔注射大肠杆菌小鼠TLR信号通路MyD88、ERK及细胞因子的影响[J].浙江农业科学,2017,58(1):143-147.LIU C,WANG C,ZUO Z C,et al.Effects of Astragalus polysaccharides on TLR signaling pathway MyD88,ERK and cytokines in mice with intraperitoneal injection of Escherichia coli[J].Journal of Zhejiang Agricultural Sciences,2017,58(1):143-147.(in Chinese)
[18] 曹利军.人参皂甙Rg1预处理对ConA诱导肝损伤的保护作用及机制研究[D].上海:第二军医大学,2013.CAO L J.The therapeutic effect and mechanisms of ginsenoside Rg1 on ConA-induced liver injury[D].Shanghai:The Second Military Medical University,2013.(in Chinese)
[19] 周恒,谢国敏,周珂,等.白介素22融合蛋白对刀豆蛋白A诱导肝损伤病理状态下肝切除后肝再生的影响[J].安徽医科大学学报,2017,52(8):1109-1114.ZHOU H,XIE G M,ZHOU K,et al.Effect of IL-22-Fc fusion protein on liver regeneration in mice after hepatectomy under ConA-induced hepatic injury condition[J].Acta Universitatis Medicinalis Anhui,2017,52(8):1109-1114.(in Chinese)
[20] 金燕,王惠萍,肖延风,等.血常规、C反应蛋白及降钙素原在新生儿败血症中的诊断价值[J].中国妇幼健康研究,2017,28(6):647-649.JIN Y,WANG H P,XIAO Y F,et al.Diagnostic value of blood routine examination,C-reactive protein and procalcitonin in neonatal sepsis[J].Chinese Journal of Maternal and Child Health Research,2017,28(6):647-649.(in Chinese)
[21] MAVROMMATIS A C,THEODORIDIS T,ORFANIDOU A,et al.Coagulation system and platelets are fully activated in uncomplicated sepsis[J].Critical Care Medicine,2000,28(2):451-457.
[22] CREMER M,WEIMANN A,SZEKESSY D,et al.Low immature platelet fraction suggests decreased megakaryopoiesis in neonates with sepsis or necrotizing enterocolitis[J].Journal of Perinatology,2013,33(8):622-626.
[23] AMéLIE R,KINDA S,SYLVIE C,et al.Severe sepsis and septic shock associated with chikungunya virus infection,Guadeloupe,2014[J].Emerging Infectious Diseases,2016,22(5):891-894.
[2] BONE R C,SIBBALD W J,SPRUNG C L.The ACCP-SCCM consensus conference on sepsis and organ failure[J].Chest,1992,101(6):1481-1483.
[3] EFFENBERGER-NEIDNICHT K,HARTMANN M.Mechanisms of hemolysis during sepsis[J].Inflammation,2018,41(5):1569-1581.
[4] 梁译丹.胡椒碱抑制ATP诱导的巨噬细胞焦亡及抗败血症的机制研究[D].广州:暨南大学,2017.LIANG Y D.The mechanism of piperine on suppressing ATP-induced pyroptosis and bacterial sepsis[D].Guangzhou:Jinan University,2017.(in Chinese)
[5] JAWAD I,LUK information on the burden of sepsis:Global estimates of incidence,prevalence and mortality[J].Journal of Global Health,2012,2(1):10404.
[6] 栾正刚.急性胰腺炎肺损伤中转录调节因子作用的实验研究[D].沈阳:中国医科大学,2003.LUAN Z G.The role of nuclear factor-kappaB activation in the lung injury of murine acute necrotizing pancreatitis[D].Shenyang:China Medical University,2003.(in Chinese)
[7] LI R W,LIN G D,MYERS S P,et al.Anti-inflammatory activity of Chinese medicinal vineplants[J].Journal of Ethnopharmacology,2003,85(1):61-67.
[8] 庞佶,郭金鹏,金敏,等.鸡血藤水提物抗流感病毒作用的研究[J].中国卫生检验杂志,2015,25(4):488-490.PANG J,GUO J P,JIN M,et al.Anti-influenza virus effects of aqueous extract from Spatholobus sub-erectus[J].Chinese Journal of Health Laboratory Technology,2015,25(4):488-490.(in Chinese)
[9] 胡庭俊,何彩美,钟晶.鸡血藤不同极性段提取物制备及体外抑菌作用初步研究[J].动物医学进展,2010,31(9):33-36.HU T J,HE C M,ZHONG J.Preparation and antibacterial activity of extracts from different polar segments of Spatholobus suberectus Dunn[J].Progress in Veterinary Medicine,2010,31(9):33-36.(in Chinese)
[10] CHEN H L,YANG J,FU Y F,et al.Effect of total flavonoids of Spatholobus suberectus Dunn on PCV2 induced oxidative stress in RAW264.7 cells[J].BMC Complementary & Alternative Medicine,2017,17(1):244.
[11] FU Y,JIANG L,ZHAO W,et al.Immunomodulatory and antioxidant effects of total flavonoids of Spatholobus suberectus Dunn on PCV2 infected mice[J].Scientific Reports,2017,7(1):8676.
[12] 陈海兰,蒙西南,赵尉丹,等.鸡血藤总黄酮乙酸乙酯部位对大肠杆菌脂多糖(LPS)刺激RAW264.7免疫细胞氧化应激的调节作用[J].江苏农业科学,2017,45(8):154-156.CHEN H L,MENG X N,ZHAO W D,et al.Regulatory effect of ethyl acetate extract of total flavonoids of Caulis spatholobi on lipopolysaccharide induced oxidative stress in RAW264.7 cells[J].Jiangsu Agricultural Sciences,2017,45(8):154-156.(in Chinese)
[13] 陈海兰,赵尉丹,付远妨,等.鸡血藤总黄酮抗炎活性的研究[J].黑龙江畜牧兽医,2017,11:211-213.CHEN H L,ZHAO W D,FU Y F,et al.The anti-inflammatory effect of total flavonoid of Spatholobus suberectus Dunn[J].Heilongjiang Animal Science and Veterinary Medicine,2017,11:211-213.(in Chinese)
[14] 付远妨,赵尉丹,蒙西南,等.鸡血藤黄酮乙酸乙酯部位对H2O2诱导RAW264.7细胞氧化应激的调节作用[J].黑龙江畜牧兽医,2017,12:145-147.FU Y F,ZHAO W D,MENG X N,et al.Regulatory effect of ethyl acetate extract of total flavonoid of Spatholobus suberectus Dunn on the oxidative stress in H2O2 stimulated RAW264.7 cells[J].Heilongjiang Animal Science and Veterinary Medicine,2017,12:145-147.(in Chinese)
[15] 韩玉杰.白介素17(IL-17)在大肠埃希菌诱导的新生大鼠败血症中的意义[D].苏州:苏州大学,2012.HAN Y J.The significance of interleukin 17 (IL-17) in neonatal rats septicemia with E.coli[D].Suzhou:Suzhou University,2012.(in Chinese)
[16] 厉世伟,谷静娟,赵璐.金银花、连翘对大肠杆菌感染模型小鼠血清Th1/Th2细胞因子的调节作用[J].现代中西医结合杂志,2013,22(6):594-596.LI S W,GU J J,ZHAO L.Regulatory mechanism of Flos lonicerae and Fructus forsythia for mouse infected by Escherichia coli[J].Modern Journal of Integrated Chinese Traditional and Western Medicine,2013,22(6):594-596.(in Chinese)
[17] 刘灿,王婵,左之才,等.黄芪多糖对腹腔注射大肠杆菌小鼠TLR信号通路MyD88、ERK及细胞因子的影响[J].浙江农业科学,2017,58(1):143-147.LIU C,WANG C,ZUO Z C,et al.Effects of Astragalus polysaccharides on TLR signaling pathway MyD88,ERK and cytokines in mice with intraperitoneal injection of Escherichia coli[J].Journal of Zhejiang Agricultural Sciences,2017,58(1):143-147.(in Chinese)
[18] 曹利军.人参皂甙Rg1预处理对ConA诱导肝损伤的保护作用及机制研究[D].上海:第二军医大学,2013.CAO L J.The therapeutic effect and mechanisms of ginsenoside Rg1 on ConA-induced liver injury[D].Shanghai:The Second Military Medical University,2013.(in Chinese)
[19] 周恒,谢国敏,周珂,等.白介素22融合蛋白对刀豆蛋白A诱导肝损伤病理状态下肝切除后肝再生的影响[J].安徽医科大学学报,2017,52(8):1109-1114.ZHOU H,XIE G M,ZHOU K,et al.Effect of IL-22-Fc fusion protein on liver regeneration in mice after hepatectomy under ConA-induced hepatic injury condition[J].Acta Universitatis Medicinalis Anhui,2017,52(8):1109-1114.(in Chinese)
[20] 金燕,王惠萍,肖延风,等.血常规、C反应蛋白及降钙素原在新生儿败血症中的诊断价值[J].中国妇幼健康研究,2017,28(6):647-649.JIN Y,WANG H P,XIAO Y F,et al.Diagnostic value of blood routine examination,C-reactive protein and procalcitonin in neonatal sepsis[J].Chinese Journal of Maternal and Child Health Research,2017,28(6):647-649.(in Chinese)
[21] MAVROMMATIS A C,THEODORIDIS T,ORFANIDOU A,et al.Coagulation system and platelets are fully activated in uncomplicated sepsis[J].Critical Care Medicine,2000,28(2):451-457.
[22] CREMER M,WEIMANN A,SZEKESSY D,et al.Low immature platelet fraction suggests decreased megakaryopoiesis in neonates with sepsis or necrotizing enterocolitis[J].Journal of Perinatology,2013,33(8):622-626.
[23] AMéLIE R,KINDA S,SYLVIE C,et al.Severe sepsis and septic shock associated with chikungunya virus infection,Guadeloupe,2014[J].Emerging Infectious Diseases,2016,22(5):891-894.