摘要
目的探讨乌司他丁(ulinastatin,UTI)在大鼠肾脏机械性损伤后对肾脏白细胞介素6(Interleukin-6,IL-6)和热休克蛋白70(Heat shock protein70,HSP70)表达的影响及意义。
方法成年雌性Wistar大鼠66只,随机分为非创伤对照组(C组)、单纯创伤组(T组)、创伤后给予乌司他丁组(U组)。后两组动物分别与创伤后1h、6h、12h、24h、36h各处死六只。采用自由落体生物撞击仪撞击大鼠脊肋角复制创伤动物模型。HE染色观察损伤后肾脏的基本病理改变。用组织芯片技术及免疫组织化学方法检测各组创伤后1 h、6 h、12 h、24 h、36 h点肾脏组织中IL-6和HSP70蛋白的表达。
结果对照组肾脏组织中IL-6和HSP70蛋白均呈弱阳性表达。与对照组比较,单纯创伤组在创伤后1、6、12、24 h点IL-6表达明显增强(P＜0.05),6 h点HSP70表达明显增强(P＜0.05),各时相点Paller评分均升高(P＜0.05)。创伤后注射乌司他丁组各时相点肾组织中IL-6表达明显低于单纯创伤组(P＜0.05),HSP70表达在1、24、36 h点明显高于单纯创伤组(以1 h点为著,P＜0.05),且创伤后各时相点Paller评分均降低(P＜0.05)。Spearman等级相关分析显示:单纯创伤组HSP70表达与IL-6表达呈正相关(P＜0.05);创伤后注射乌司他丁组HSP70表达与IL-6表达呈负相关(P＜0.01)。
结论1、急性创伤后早期肾脏IL-6的表达水平反映了肾小管上皮细胞的损伤及其炎性反应程度。2、IL-6在调节炎症方面具有双重功能,在发挥炎性损伤作用的同时,又通过其炎性刺激作用诱导肾小管上皮细胞表达HSP70发挥抗炎作用。3、在无外源性药物干预的情况下,机体自身经抗炎作用而产生的HSP70,在单纯性创伤早期没有对肾脏起到有效的保护作用。4、急性创伤后12小时至36小时,乌司他丁通过上调肾小管上皮细胞HSP70的再次高表达以保护肾脏的缺血再灌注损伤。5、乌司他丁主要在创伤后早期(以1小时为著)发挥作用,通过抑制肾小管上皮细胞IL-6的过度表达,上调HSP70的表达,纠正两者表达水平之间的比例失衡,而对大鼠肾脏起到有效的保护作用。
OBJECTIVE To investigate the significance of ulinastatin's effects on expression of IL-6 and HSP70 in kidney after mechanical injury.
METHODS Sixity-six adult female Wistar rats were randomly divided into three groups:control(C,n=6),trauma(T,n=30),and injected ulinastatin after trauma(U,n=30).The 6 rats in the last two groups were excuted at 1h,6h,12h,24h and 36h after trauma respectively.The model of trauma rats was made with free-fall bio-impactor striking the spinal column and ribs of rats.Renal tissue was observed after HE stained. The expression of IL-6 and HSP70 in each group's renal tissue were detected with tissue chip and immunohistochemical stain at 1,6,12,24 and 36 h after trauma.
RESULTS The expression of IL-6 and HSP70 in renal tissue was low in control group.Compared with control,the expression of IL-6 in trauma group was markedly increased at 1,6,12,24 h after trauma(P＜0.05),the expression of HSP70 was significantly increased at 6h after trauma (P＜0.05),injury score heightend correspondly(P＜0.05).The expression of IL-6 in group injected Ulinastatin after trauma were more lower than trauma group,the expression of HSP70 at 1,24,36 h after trauma was higher than trauma group especially at 1 h(P＜0.05),meanwhile,injury score decreased correspondly(P＜0.05).The Spearman correlation analyse revealed that in group T the expression between HSP70 and IL-6 present positive correlation(P＜0.05),but in group U the expression between HSP70 and IL-6 present negative correlation(P＜0.01).
CONCLUSION 1.IL-6 expression at early times after trauma can reflex the level of injury and inflammatory reaction of renal tissue. 2.IL-6 have dual effects on adjust inflammation.It not only lead to the inflammatory injury,but also induce HSP70 expression of renal tubular epithelial cell to produce anti-inflammatory effects through its inflammatory excitation.3.Without exogenous medicine's intervention, the HSP70 produced by organism itself have not play a effective protection part in against inflammatory injury at early times after trauma.4.From 12h to 36h after trauma,Ulinastatin can reduce the risk of Ischemia reperfusion injury through up-regulation of HSP70 expression. 5.Ulinastatin can reduce the risk of renal injury at early times after trauma especially at 1 hour.The protective effects of ulinastatin associated with the decrease of IL-6 expression,up-regulation of HSP70 expression,and recitify the unbalanced proportion between HSP70 and IL-6 expression.
引文
[1]Peden M,Scurfield R,Sleet D,et al.World report on road traffic injury prevention[R].Geneva:WHO,2004:3229.
[2]杨唐俊,何斌.肾脏损伤.见:王正国 主编.创伤病理学.第1版,上海,上海科学技术出版社,2002;1562-1564.
[3]秦荣良,等.闭合性肾损伤的诊断和外科治疗.中华外科杂志,1996;34:505.
[4]Hiroshi Ogura,Hiroshi Tanaka,Taichin Koh,et all Priming,second -hit priming,and apoptosis in leukocytes from trauma patients[]]J Trauma,1999,46(5):774-783
[5]Ono S,Ichikura T,Mochizuki H.The pathogenesis of the systemic inflammatory response syndrome and compensatory antiinflammatory response syndrome following surgical stress[J].Nippon Geka Gakkai Zasshi,2003,104:499-505.
[6]陈依君.乌司他丁对机体的保护作用.上海第二医科大学学报,2005;25.863-864
[7]景炳文.乌司他丁在急危重症临床应用的进展[J].中国危重病急救医学,2006:18(12):117-120。
[8]邓文锋,于立新,余玉明,等.乌司他丁减轻移植肾热缺血损伤的临床研究[J].中华泌尿外科杂志,2004,25:739-741.
[9]李昕,刘佳佳.IL-6对中性粒细胞在炎症中作用的影响[J].国外医学免疫分册,2005;28(5):277-280.
[10]Damas P,Canivet J,Groo re,et al.Sep sis and serum cytok ine concentrations[]].Crit Care M ed,1997,25:405-412.
[11]Youker K,Smith CW,Ander son DC,et al.Neutrophil adherence to isolated adult cardiac myocytes.Induction by cardiac lymph collected during ischemia and reperfusion.J Clin Invest, 1992,89(2):602
[12]Pape HC,van Griensven M,Rice J,et al.Major secondary surgery in blunt trauma patients and perioperative cytokine liberation:determination of the clinical relevance of biochemical markers.J Trauma.2001,50(6):989
[13]唐昱.热休克蛋白70与心肌保护的研究进展[J].国际检验医学杂志,2006;27(11):1028-1030
[14]Lepore DA,Knight KR,Anderson RL,et al.Role of Priming Stresses and HSPTO in protection from ischemia-reperfusion injury in cardiac and skeletal muscle.Cell Stress Chaperones,2007,6:93-96.
[15]孙玲娣,赵浦,等.HSP70在缺血再灌注大鼠大鼠肾脏中的表达及意义[J].中国小儿急救医学,2006;13(4):349-351。
[16]Tano naka K,Toga W,Takeo S.Induction of heat shock protein 70in failing heart.Nippon Yakurigaku Zasshi,2004,123(2):71-76.
[17]Li G,Bea S,Zhang L.Effect of prenatal hypoxia on heat stressmediated cardioprotection in adult rat heart[J]Am J Physiol Heart Circ Physiol,2004,286(5):H1712-1719.
[18]蒙伶俐,孙少华,等.经体表创伤后肾脏组织中缺氧诱导因子-1 α表达的调控[J].第四军医大学学报,2005;26(14):1311-1314.
[19]SmithJK kidney trauma[EB/OL].http://www.emedicine,com/radio/topic376html,2004-5-17/2004-5-29.
[20]Paller MS.Oxygen free radicals in ischemic acute renal failure in the rat[J].Clin Invest 1998;74:1156.
[21]Krran MC,Smith MD,Weedon H et al Measurement of cytokine and adhesion molecule expression in synovial tissue by digital image analysis.Ann Rheum Dis,2001;60;296-298.
[22]Sconocchia G,Campagnano L,Adorno D,et al CD44 ligation on peripheral blood polymorphonuclear cells induces interleukin26production[J].Blood,2001,97(11):3621-3627
[23]Riedemann NC,Guo RF,Hollmann TJ,et al.Regulatory role of C5a in LPS-induced IL-6 production by neutrophils during sepsis[J]FASEB J,2004,18(2):370-372
[24]Xing Z,Gauldie J,Cox G,et al.IL-6 is an antiinflammatory cytokine required for controlling local or systemic acute inflammatory responses[J].J Clin Invest,1998,i01(2):311-312
[25]Steensberg A,Fischer CP,Keller C,et al.IL-6 enhances plasma IL-Ira,IL-10,and cortisol in humans[J]Am J Physiol Endocrinol Metab,2003,285(2):E433-E437.
[26]Rabergh CM,Airaksinen S,Soitamo A,et al.Tissue specific expression of zebrafish(Danio rerio)heat shock factor ImRNAs in response to heat stress[J].Journal of Experimental Biology,2000,203(12):177-181
[27]Laszlo A.Evidence for two states of thermotolerance in mammaliancells[J].Int J Hyperthermia,1998,4:513.
[28]Riabowol K T,Mizzen L A,Welch WJ.Heat shock is lethal to fibroblasts microinjected ptorein[J].Cell,1986,46:959-961.
[29]Hotch kiss R,Nunnally I,Lindquist S.Hyperthermia protects mice against the lethal effects of endotoxin[J].Am J Physiol,1993,265:R1447-1457.
[30]Currie RW,Tanguay RW.Heat shock response and.limitation of tissue necrosis during occlusion/ reperfusion in rabbit hearts[J]Circulation,1993,87:963-971.
[31]李立,田大光,李晓研.等.腹部大手术后运用乌司他丁减少手术侵袭的临床观察[J].中华肝胆外科杂志,2001,7:315-316.
[32]Nouman J.The role of cytokine in the pathogenesis of acute pancrea -titis[J].Am J Surg,1998,175:76-83.
[33]Sato N,Endo S,Kimura Y,et al.Influence of a human proteasein.hibitor on surgical stress induced immunOsuppressiOn[J].Dig Surg,2002,19(4):300-305.
[34]Aosasa S,Ono S,Mochizuki H,et al.Mechanism of the inhibitory efect of protease inhibitor on tumor necrosis factor alpha production of monocytes[J].Shock,2001,15(2):101-105.
[35]Tani T,Abe H,Endo Y,et al.Efect of a urinary trypsin inhibitor on acute circulatory insufficiency after surgical operation[J].Am J Surg,1998,175(2):142-145.
[36]Komori M,Takada K,Tomizawa Y,et al.Urinary trysin inhibitor improve peripheral microc irculation and bronchospasm associated with systemic anaphylaxis in rabbits vivo[J].Shock,2003,20(2):189-194.
[37]Takada K,Komori M,Notoya A.et al.Efect of ulinastatin on mi.crocirculation during excessive hemorrhage using fluid therapy[J].in vivo,2003,17(2):129-135.
[38]Tale S,Yokono S,Ueki M.et al.Efects of ulinastatin(urinary trypsin inhibitor)on ATP,intracellular Ph,and intracellular sodium transients during ischemia and reperfusion in the rat kindney in vivo[J].J Anesth,2001,15(1):33-38.
[39]金周慧,等.细胞因子网络在肾脏疾病中的作用[J].中国实用内科杂志,2001;21(9):562-564。
[40]陈婷婷,王刚,等.蛋白酶抑制剂乌司他丁对血管内源性活性物质的影响[J].第四军医大学学报,2005;26(3):220-222.
[1]Mat sumi M,Mizobuchi S,Kaku R,et al.Changes of urinary t rip sin inhibitor in blood and urine,as well as serum cytokines in living related liver t ransplantation[J].Masui,2003,52(3):251-256.
[2]尹勇,高乃荣,李兆丽.乌司他丁对急性坏死性胰腺炎大鼠肺损伤的保护作用[J].世界华人消化杂志,2002,10(5):558-561。
[3]吴稚莉,海燕,田炳章.乌司他丁对急性坏死性胰腺炎模型大鼠血清淀粉酶、肿瘤坏死因子的影响[J].中国药房,2004,15(9):535-536.
[4]上海乌司他丁临床试验协作组.国产乌司他丁治疗急性胰腺炎的多中心临床研究[J].中华消化杂志,2001,21(5):293-296。
[5]王刚,陈婷婷,高长青.乌司他丁对创伤失血性休克兔肺损伤的保护作用[J].中国危重病急救医学,2005,17(1):36-38。
[6]Sugita T,Watarida S,Kat suyama K,et al.Effect of a human urinary protease inhibitor(Ulinastatin)on respiratory function in pediatric patient s undergoing eardiopulmonary bypass[J].J Cardiovase S urg(Torino),2002,43(4):437-440.
[7]Furukawa K,Kobayashi R,Kuroda K,et al.A case of severe alcoholic hepatitis with successfully t reared by the therapy for hypereytokinemia and granulocytic hyperelastasemia[J].Nippon S hokakibyo Gakkai Zasshi,2002,99(4):401-405.
[8]Kobayashi H,Suzuki M,Sun GW,et al.Suppression of urokinasetype plasminogen activator expression from human ovarian cancer cells by urinary trypsin inhibitor[J].Bioehim Bophys A cta,2000,1481(2):310-316.
[9]Kobayashi H,Shinohara H,Gotoh J,el al.Antimetastanic therapy by Urinary trypsin inhibitor in combination with an antieaneer agent [J].Br J Cancer,1995,72(5):1131-1137.
[10]钱叶本,赵红川,熊奇如,等.乌司他丁在肝脏缺血再灌损伤和肝脏再生中的作用[J].中华肝胆外科杂志,2001,11(7):701-702.
[11]于金贵,周广利,孟冬梅,等.乌司他丁对心脏直视手术患者体外循环期炎性细胞因子和自由基的影响[J].中华麻醉学杂志,2001,21(12):724-726。
[12]李伟,王剑,蒋艳波,等.乌司他丁对术中单肺通气患者白介素-8的影响[J].癌症,2003,22(10):1074-1076。
[13]Hiyama A,Takeda J,Kotate Y,et al.A human urinary protease inhibitor(ulinastatin)inhibit s neut rophilext racellular release of elastase during cardiopulmonary bypass[J].Cardiothorac Vasc A nesth,1997,11:580-584.
[14]邵明义,张良清,邓烈华,等.乌司他丁对全身炎症反应综合征的治疗作用[J].中国危重病急救医学,2005,17(4):228-230.
[15]Yamada S,Takat suka H,Takemoto Y,et al.Urinary trypsin inhibitor concent ration can predict the immunological insult of chemotherapy and complications af terbone marrow t ransplantation[J].Bone Marrow Transplant,2001,27(2):195-199.
[16]胡珏,陆小华,周益锋.乌司他丁对全髋置换患者手术期间免疫功能的影响[J].中华医药杂志,2005,5(2):109-110。
[17]迟达超,韩希望,间立昆,等.乌司他丁对胃肠癌手术患者免疫功能的影呐[J].第四军医大学学报,2004,25(15):1435-1437.
[18]Sato N,Endo S,Kimura Y,et al.Influence of a human protease inhibitor on surgical st ress induced immunosuppression[J]Di g S urg,2002,19(4):300-305.
[19]杨振林,郑树森,朱峰,等.两种不同途径应用乌司他丁治疗重症急性胰腺炎的疗效观察.中华肝胆外科杂志,2005,11(2):101-103。
[20]Masuda T,Sato K,Noda C,et al.Protective efect of urinary trypsin inhibitor on myocardial mitochondria during hemorrhagic shock and reperfusion.Crit Care Med,2003,31(7):1987-1992
[21]陈聪聪,柳子明,王慧华,等乌司他丁对大鼠肾缺血/再灌注损伤的保护作用.中华麻醉学杂志,2004,24(11):828-832.
[22]李颖川,姜桢.尿蛋白酶抑制剂防治肺缺血再灌注损伤的实验研究.复旦 学报(医学版),2004,31(2):182-185.
[23]谢康,黄跃生.乌司他丁对严重烧伤大鼠血清中致炎因子的影响及意义.第三军医大学学报,2004,26(18):1643-1645。
[24]任学群,傅侃达,万相斌,等.乌司他丁和全肠外营养对胃肠道恶性肿瘤患者术后急性相蛋白代谢的影响.中华普通外科杂志,2003,18(5):287-289.
[25]Sato A,Kuwabara Y,Shinoda N,et al.Use of low dose dopamine,gabexate mesilate and ulinastatin roduces the water balance and pulmonary omphcation in thoracic esophagectomy patients.Dis Esophagus,2005,18(3):151-154.
[26]李国强,王学浩,孙倍成,等.乌司他丁对肝切除患者围手术期免疫功能的影响.中华肝胆外科杂志,2002,8(1):120-122.
[27]卢中秋,赵初环,李惠萍,等.抗菌药物联合乌司他丁对酒精性肝病大鼠创伤弧菌脓毒症的作用.中华急诊医学杂志,2005,14(6):474-478.
[28]Inoue K,Takan o H,Shimada A,et at.Urinary trypsin inhibitor protects against systemic inflammation induced by lipopolysaccharide.Mol Pharmacol,2005,67(3):673-680.
[29]谭黎杰,薛亮,徐正浪,等.乌司他丁治疗同种大鼠单肺移植后缺血再灌注损伤.中华外科杂志,2005,43(14):934-935.
[30]宋一民,宋春芳,许评,等.乌司他丁对犬胰岛的保护作用.中华器官移植杂志,2005,26(6):353-355
[31]Yu JR,Yah S,Liu XS,et al.Attenuation of graft ischemia-repedusion injury by urinary trypsin inhibitor in mouse intestinal transplantation.World J Gasta'oenteml,2005,11(11):1605-1609.
[32]张欢,乔青,杨拔贤,等.乌司他丁对原位肝移植术患者围术期促炎性细胞因子和氧自由基代谢的影响.中华麻醉学杂志,2003,23(5);325-327。
[33]余剑波,姚尚龙,袁世荧.心内直视手术中应用乌司他丁对β-GCD、Gel 及Fn的影响.中华麻醉学杂志,2004,24(3):165-167。
[34]徐康清,孙培吾,黄文起,等.乌司他丁对体外循环心脏手术后肺功能的影响.中华胸心血管外科杂志,2004,20(1).7-9。
[35]何小京,王亚平,常业恬,等.乌司他丁对体外循环期间胃肠循环及炎性因子的影响.中华麻醉学杂志,2003,23(4):253-256.
