缺血再灌流肾组织内皮素—1及其受体基因表达的实验研究

为了探究内皮素１（ＥＴ１）对肾功能的影响和作用方式，采用斑点杂交和原位杂交方法对大鼠缺血６０分钟再灌注肾组织ＥＴ１及其受体亚型（ＥＴＡ、ＥＴＢ）的基因表达进行了研究。结果发现：再灌流１小时，ＥＴ１、ＥＴＡ、ＥＴＢｍＲＮＡ均明显升高；再灌流２４小时仍维持较高水平。ＥＴ１和ＥＴＡｍＲＮＡ杂交信号再灌流３小时达高峰。ＥＴ１ｍＲＮＡ主要分布肾皮质小血管内皮细胞、髓质肾小管和集合管，ＥＴＡ受体ｍＲＮＡ则分布于上述小血管的平滑肌细胞。ＥＴＢ受体ｍＲＮＡ于再灌流６小时达高峰，主要分布髓质肾小管、集合管。说明缺血再灌流肾内皮素受体亚型上调在皮质以ＥＴＡ为主，在髓质以ＥＴＢ为主，分别与增强表达的ＥＴ１结合导致肾皮质缺血和水钠代谢异常。     

内皮素在急性肾缺血再灌注损伤中作用的实验研究

内皮素在急性肾缺血再灌注损伤中作用的实验研究涂响安余明年谢佛龙赵志毅冯家骅何炳辉彭轼平用放射免疫方法（ＲＩＡ）检测大鼠左肾动脉夹闭６０分钟致急性肾缺血再灌注损伤（ＡＲＲＩ）模型其血浆和肾组织内皮素（ＥＴ）水平变化，以探讨ＥＴ在急性肾缺血再灌注损伤中的...     

内皮素-1与肝脏缺血再灌注损伤的实验研究

目的：探索内皮素－1（ET－1）在肝脏血再灌注损伤中的作用。方法：选择雄性Wistar大鼠80只,分为正常对照组、缺血再灌注组1生理盐水组和ET－1抗体组、观察肝脏缺血60min再灌注3h后血浆ET－1、丙氨酸转氨酶（ALT）、透明质本酸（HA）、以及肝组织中ET－1和丙二醛（MDA）含量的变化,并观察肝组织病理学变化,同时,在缺血再灌注组选择第1、3、6、12和24h时相点观察ET－1的变化规律。结果：肝脏缺血再灌注后,血浆和肝组织中ET－1,血浆HA`ALT肝组织中MDA显著升高,而ET－1抗体组血浆ET－1、HA、ALT与缺血再灌注组相比显著降低（P＜0．01,P＜0．05）,同时,肝组织的瘀血程度和损伤程度显著改善。结论ET－1参与了肝脏缺血再灌注损伤,这种损伤与肝脏微循环障碍有关。     

抗氧化剂防治肾缺血再灌流损伤的研究

抗氧化剂防治肾缺血再灌流损伤的研究吴雄飞，李为兵，帅学焱，金锡御氧自由基在组织缺血再灌流损伤中起重要作用，已由实验证实并日益引起重视。我们在家兔肾动脉钳夹致肾缺血再灌流模型上，采用腹主动脉灌注直接给药的方法，观察抗氧化剂超氧化物歧化酶（ＳＯＤ）；过氧...     

内皮素和降钙素基因相关肽在急性肾缺血再灌注损伤中的作用

为研究大鼠急性肾缺血再灌注损伤血液和肾组织中内皮素（ＥＴ）和降钙素基因相关肽（ＣＧＲＰ）的动态变化规律，放射免疫法检测急性肾缺血再灌注损伤大鼠血浆和肾皮质、髓质ＥＴ和ＣＧＲＰ水平变化，结果肾组织ＥＴ水平于再灌注３ｈ达到高峰，２４ｈ仍维持较高水平，肾组织ＣＧＲＰ水平再灌注２４ｈ达到高峰，血浆ＥＴ和ＣＧＲＰ于再灌注损伤后虽升高，但无显著性差异，认为肾组织ＥＴ和ＣＧＲＰ升高在急性肾缺血再灌注损伤和修复中     

内皮素-1单克隆抗体对肝移植缺血再灌注损伤保护作用的实验研究

目的: 探讨内皮素-1(ET-1)抗体对移植肝缺血再灌注损伤的保护作用.方法: 选用雄性SD大鼠108只,分为对照组、肝移植组与ET-1抗体组,观察应用抗ET-1单抗前后移植肝热缺血15 min,冷缺血40 min再灌注后第4小时血浆ET-1、丙氨酸转移酶(ALT)、透明质酸(HA)以及肝组织中ET-1和丙二醛(MDA)含量及肝组织病理学变化;同时,在移植肝再灌注后的第1,4,12,24小时观察ET-1的变化规律.结果: 移植肝缺血再灌注后,血浆和肝组织中ET-1,血浆HA,ALT和肝组织中MDA显著升高,而ET-1抗体组血浆和肝组织中ET-1,血浆HA,ALT和肝组织中MDA与肝移植组相比显著降低(P<0.01,P<0.05 ),且肝组织的淤血程度和损伤程度显著改善.结论: ET-1单克隆抗体对移植肝缺血再灌注损伤有保护作用.     

内皮素-1单克隆抗体对肝移植缺血再灌注损伤的保护作用

缺血再灌注损伤是导致移植肝无功能的常见原因之一。我们应用ET 1单克隆抗体灌注移植肝 ,研究了ET 1单抗在肝移植缺血再灌注损伤中的作用。一、材料与方法1.材料 :实验所用SD大鼠由中山大学实验动物中心提供 ,全部为雄性 ,体重2 2 0～ 2 5 0 g。内皮素 1(ET 1)单克隆抗体及试剂盒购自深圳晶美公司 ;透明质酸 (HA)试剂盒购自海军医学研究所 ;丙二醛 (MDA)试剂盒由南京建成生物工程研究所提供 ;谷丙转氨酶 (ALT)检测由中山大学附属一院检验科协助完成。2 .肝移植动物模型制作及分组 :按Kamada等[1] 的方法行双袖套法大鼠原位肝移植术 …     

葛根素对家兔肝缺血再灌注一氧化氮、内皮素-1的影响

目的观察一氧化氮(NO)、内皮素-1(ET-1)在家兔肝缺血再灌注损伤过程中的变化及葛根素(Pur)对其的影响。方法实验家兔随机分为3组:对照组(C组,n=10)、缺血再灌注组(IR组,n=10)和葛根素治疗组(Pur组,n=10)。分别测定缺血前,缺血第45分钟及再灌注第45分钟时肝组织NO,ET-1含量,用电镜观察家兔HIRI及经Pur保护后的肝组织形态学改变。结果家兔缺血再灌注第45分钟时,与C组、Pur组比较:IR组血浆及组织NO含量均明显较低(P<0.01,P<0.05);IR组血浆及组织ET-1含量明显较高(P<0.01,P<0.05);与IR组比较,Pur组肝组织超微结构明显改善。结论葛根素可通过调节机体NO和ET-1水平而对肝缺血再灌注损伤发挥积极的保护作用。     

丙泊酚对大鼠肝脏缺血-再灌注内皮素-1和肝功能的影响

目的 探讨丙泊酚对肝脏缺血-再灌注损伤中内皮素-1(ET-1及肝功能的影响.方法 SD大鼠72只,体重280～300 g,随机均分为三组:缺血-再灌注组(Ⅰ组)、丙泊酚保护组(Ⅱ组)、对照组(Ⅲ组).在肝脏缺血-再灌注前(T1)、再灌注后2 h(T2)、4 h(T3)时检测血浆和肝组织中ET-1、谷丙转氨酶(ALT)及肝脏组织病理学变化.结果 T2、T3时Ⅰ、Ⅱ组血浆、肝组织中ET-1水平明显高于Ⅲ组(P＜0.05),而Ⅱ组明显低于Ⅰ组(P＜0.05).T2、T3时Ⅰ、Ⅱ组血浆、肝组织中ET-1水平、血浆ALT明显高于T1时,而Ⅰ组T3时高于T2时(P＜0.05).结论 ET-1参与了肝脏缺血再灌注损伤,丙泊酚具有抑制ET-1分泌及保护肝功能的作用.     

血红素氧合酶-1对大鼠肾缺血再灌注损伤的保护作用

目的　探讨钴卟啉 (CoPP)诱导的血红素氧合酶 (HO) 1高表达对大鼠肾缺血再灌注损伤 (IRI)的保护作用。方法　建立大鼠肾缺血再灌注损伤模型 ,随机将动物分为假手术组 ,对照组和实验组。动态检测血尿素氮 (BUN)、肌酐 (Cr)、超氧化物岐化酶 (SOD)、丙二醛 (MDA)的含量以及进行肾组织光镜形态学观察和酶联免疫吸附试验 (ELISA)和Westernblot分析HO 1。结果　对照组BUN、Cr升高 ,SOD下降 ,MDA升高 ,HO 1中度提高 (14 4.5± 13 .6) ,肾组织结构紊乱。实验组除HO 1含量大幅提高外 (62 9.4± 78.9) ,尚能显著逆转上述改变 ,两组间差异具有统计学意义 (P <0 .0 5 )。结论　CoPP预处理诱导HO 1在肾缺血之前高表达可通过清除氧自由基 (OFRs)而减轻大鼠肾IRI。     

大鼠肾冷缺血再灌注损伤模型的建立

目的 建立大鼠肾冷缺血再灌注损伤(IRI)的模型.方法 封闭群SD大鼠24只,随机分为2组(n=12):A组(对照组),B组(实验组).A组切除右肾并游离左肾蒂,60 min后关闭腹腔切口.B组采用冷缺血再灌注模型,主要步骤:(1)冷灌注:右肾动脉插管对左肾原位灌注.通过右肾静脉插管将灌注液引流出体外,完成冷灌注后切除右肾,阻断左肾蒂.(2)冷缺血保存:将已充分游离的左肾牵至腹腔外,在自制保存袋中冷保存.(3)再灌注:60min后,去除保存袋,开放血流,再灌注左肾,左肾复位,缝合切口;2组大鼠均在术后24 h再次手术切除左.肾.肾组织进行光镜、电镜形态学检查,检测肾组织匀浆中超氧化物歧化酶(SOD)活力、丙二醛(MDA)含量,术前与术后24 h取血标本进行测定血尿素氮(BUN)、肌酐(Cr)评估肾功能.结果 (1)形态学检查(光镜与电镜超微结构):A组肾脏组织形态结构正常,B组损伤表现明显;(2)A组手术前后比较血浆BUN、Cr测定值差异均无统计学意义(P＞0.05).IR后的B组均高于术前,差异有统计学意义(P＜0.05);(3)IRI后A组肾组织匀浆SOD活力高于B组(P＜0.05),A组肾组织匀浆MDA含量测定值低于B组,差异有统计学意义(P＜0.05).结论 建立的模型要求条件简单、易行,可用于肾移植冷缺血再灌注损伤相关的研究;

Abstract：

Objective In this study,for studying IRI in kidney transplantation. ,we established the models of cold ischemia and reperfusion injury in rats. Methods Twenty four SD rats were randomly assigned to two groups:control (A) ,and experimental (B) group. Group A was only removed the right kidney. Cold ischemia reperfusion was performed as the follow-listed model in Group B. The main process of the model: ( 1) Perfusing left kidney: after resected the right kidney of the rat, one pipe was put in the remainder right renal artery to perfuse the left kidney. The perfusion flowed out through another pipe in the right renal vein. The blood vessels of left kidney were clipped after cold perfusion. (2) Cold ischemic conservancy : the operation table was leant to left side, and the left kidney was taken out of abdominal cavity then stored in a cold bag which was full of ice and water,but the vessels of that were intact. (3) Reperfusing left kidney: after 60 minutes, the clip was removed. Left kidneys of all rats in two groups were removed to be detected. Structure of the kidney was evaluated by light microscopy and electronic microscopy. Superoxide dismutase ( SOD) activity and malondialdehyde ( MDA) content in the renal tissues was examined,and the renal function was also assessed by determining the levels of blood urea nitrogen ( BUN) and serum creatinine (CR) before and 24 hours after operation. Results (1) Morphologic change (hematoxylin-eosin staining) :A normal morphology was observed by light microscopy and electon microscopy in group A.Significant injury was detected in group B. (2 ) In group A, there was not significant difference about BUN and CR between before and after operation (P ＞0. 05) ,but in Group B,those increased significantly at 24 hour after operation (P ＜0. 05). (3) Activity of SOD in renal tissues in group A was higher than those in group B (P ＜ 0. 05 ) , meanwhile, Content of MDA in group A was lower than those in group B ( P ＜0. 05 ).Conclusion The rat renal cold ischemia reperfusion model we established is feasible regardless of experimental conditions, and can be studied as the events following IRI in kidney transplantation.     

Involvement of endothelin/ nitric oxide balance in hepatic ischemia/ reperfusion injury

Introduction: Endothelin (ET) and nitric oxide (NO) act as opponents in the regulation of the hepatic microcirculation. During ischemia/reperfusion (I/R) ET levels are increased, whereas no rise in NO levels is observed. This imbalance may be responsible for microcirculatory disturbances. The aim of this study was to restore the delicate ET/NO balance to maintain the integrity of the hepatic microcirculation and to reduce I/R injury. Methods: Ischemia was induced by crossclamping of the hepatoduodenal ligament for 30 min with portal decompression using a splenocaval shunt (56 Wistar rats, 200–250 g). Sham operation, ischemia and treatment groups with the endothelin receptor antagonist (ERA) bosentan (1 mg/kg body weight i.v.) and the NO donor l-arginine (400 mg/kg body weight i.v.) were performed. For assessment of the microcirculation, sinusoidal perfusion rate, diameters of sinusoids and postsinusoidal venules, leukocyte endothelium interactions and velocity of free-flowing leukocytes were investigated by means of in vivo microscopy 30–90 min after reperfusion. Local hepatic tissue pO₂ was measured prior to ischemia, 30 min and 60 min after reperfusion and aspartate aminotransferase (AST)/alanine aminotransferase (ALT) levels were investigated 2 h and 6 h after reperfusion. Results: After ischemia, sinusoidal and venular diameters were reduced to 76% and 85%, respectively, of sham operation group values (P<0.05), but were maintained at baseline in ERA (98/102%) and NO (102/105%) groups (P<0.05). Increased postischemic leukocyte sticking in sinusoids (144%) and venules (435%) was reduced by therapy to 110/253% (ERA) and 111/ 324% (NO), respectively (P<0.05). Perfusion rate was increased to 93% and 94% compared with 82% in the ischemia group (P<0.05). Concomitant with the improved microcirculation in therapy groups, local hepatic tissue pO₂ was improved 30 min after reperfusion in the ERA (11.0 mmHg) and the l-arginine group (11.5 mmHg) relative to the ischemic group (6.9 mmHg) (P<0.05). In addition, postischemic AST/ALT increase was reduced by therapy. Conclusion: Our results indicate that maintenance of ET/NO balance by blocking ET receptors, as well as providing a NO donor, protects the liver microcirculation and reduces hepatic I/R injury. Received: 14 August 1998 Accepted: 13 October 1998     

Changes in regional renal perfusion following ischemia/reperfusion injury to the rat kidney

Summary Post-ischemic renal failure is associated with a zone of vascular hyperaemia in the outer medulla of the kidney. The effect of this lesion on regional renal perfusion is, however, unclear. Acute unilateral renal ischemia was applied to four groups of ten adult male Wistar rats for a period of 60 min, followed by revascularisation for 0, 15, 30 or 60 min. The aorta was then clamped and Microfil was injected at a standard pressure to fill the renal vasculature. Gross and histological examinations of the renal parenchyma and vasculature were then performed. Regional renal Microfil perfusion was quantified by examination of unstained histological sections, giving rise to a vascular perfusion index (VPI) for each vascular region of the kidney. The VPIs were similar in control and ischemic kidneys that were not subjected to reflow (group 1). In contrast, the VPI was markedly decreased in the inner stripe and inner medulla in animals in which revascularisation had occurred (groups 2–4), and the vasculature in these regions was histologically shown to be packed with red blood cells. Post-ischemic renal failure is associated with hyperperfusion of the medulla resulting from blockage of the vasculature that occurs during revascularisation.     

上调intermedin表达对大鼠肾脏缺血再灌注的保护作用

目的 观察上调肾脏intermedin( IMD)表达对大鼠肾脏缺血再灌注(I/R)的影响.方法 24只健康雄性Wistar大鼠随机分为假手术组、肾脏I/R组、IMD+I/R组、空质粒+I/R组,每组6只.所有动物于I/R术24h后杀检,取肾组织进行光镜检查,留取血清测定尿素氮(BUN)和血清肌酐(Scr)的浓度.免疫组织化学方法、半定量RT-PCR、Western印迹检测肾组织IMD表达及部位.Western印迹测定内皮素1(ET-1)、肿瘤坏死因子α(TNF-α)蛋白的表达.结果 HE、PAS染色结果显示,I/R组肾小管及间质病理损伤显著重于假手术组,IMD+I/R组小管间质损伤程度较肾脏I/R模型组及空质粒+I/R模型组明显减轻(1.5±0.8比7.6±2.3和7.0±1.8,均P＜0.05].与假手术组[(BUN 3.85±0.21 mmol/L,Scr(48.67±3.61) μmol/L相比,I/R组、IMD+I/R组以及空质粒+I/R组BUN(10.13±2.14) mmol/L,( 7.73±1.03) mmol/L,( 9.77±1.92) mmol/L和Scr(80.33±7.15) μmol/L,(58.50±:3.27)μmol/L,(75.67±7.58) μmol/L均明显升高(均P＜ 0.05),其中IMD+I/R组较I/R组以及空质粒+I/R组BUN和Scr水平显著降低(均P＜ 0.05).免疫组化结果显示,假手术组IMD呈弱阳性表达,主要位于肾小管间质细胞胞质内,I/R组肾组织IMD在肾小管上皮细胞和间质表达较假手术组上调;IMD+I/R组肾组织中IMD表达较I/R组明显上调(P＜0.01).与I/R组及空质粒+I/R组相比IMD+I/R组肾组织IMD mRNA,相对含量分别增加了60.7％、66.1％,蛋白相对含量分别增加了51.4％、55.9％.此外,与I/R组相比,IMD+I/R组肾组织ET-1、TNF-α蛋白表达显著降低(ET-1:0.17±0.02比0.08±0.02;TNF-α:0.35±0.02比0.21±0.04,均P＜0.05).结论 在大鼠肾脏I/R前上调IMD表达可能通过抑制ET-1、TNF-α表达保护肾脏结构及功能.     

BQ123对肺移植早期缺血再灌注损伤保护作用的实验研究

目的　研究内源性内皮素（ Ｅ Ｔ）１ 对肺移植早期缺血再灌注损伤的影响及 Ｅ Ｔ Ａ 受体阻断剂 Ｂ Ｑ１２３ 对其病理过程的保护作用。方法　以家犬（ 保存８ 小时） 左肺移植模型观察缺血再灌注。损伤过程中内源性 Ｅ Ｔ１ 产生及 Ｂ Ｑ１２３ 对其血流动力学、肺功能的作用。结果　 Ｂ Ｑ１２３ 组和对照组的平均动脉压、左房压、中心静脉压及组织形态学差异无显著性; Ｂ Ｑ１２３ 组平均肺动脉压、肺血管阻力指数及血清 Ｅ Ｔ１ 浓度明显低于对照组（ Ｐ ＜ ０ ．０１） ,心脏指数、动脉血氧分压明显高于对照组（ Ｐ ＜ ０ ．０１） ,而动脉二氧化碳分压明显低于对照组（ Ｐ ＜ ０ ．０１） ; Ｂ Ｑ１２３ 组肺组织含水百分比明显低于对照组（ Ｐ ＜０ ．０１） 。结论　 Ｂ Ｑ１２３ 对肺移植早期出现的缺血再灌注损伤具有保护作用。     

缺血再灌注损伤后小鼠肾脏血管内皮生长因子C的表达

目的探讨小鼠肾缺血再灌注损伤模型中血管内皮生长因子C的表达变化及其意义。方法建立小鼠肾缺血再灌注损伤模型。雄性C57BL／6小鼠用无创性动脉夹夹闭左肾动脉,置于32℃温箱后1h松开血管夹,取出右肾。Sham组操作同上,但不夹闭左肾动脉。再灌注0,6,12,24,48h后处死小鼠,收集外周血及肾脏标本。测定血肌酐（SCr）和尿素氮（BUN）水平。HE染色观察Sham组和缺血再灌注24h组（IR24h组）肾脏病理学变化,免疫组织化学检测Sham组和IR24h组血管内皮生长因子C（vascularendothelialgrowthfactorC,VEGF-C）在肾脏的表达及分布,连续切片检测VEGF-C与其受体血管内皮生长因子受体3（VEGFR-3）的共定位。Westernblot检测缺血后不同灌注时间VEGF-C的表达变化。结果小鼠肾脏缺血再灌注损伤后,SCr和BUN水平上升,且随着再灌注时间的延长肾功能损伤逐渐加重,再灌注24h时肾功能损伤最明显,再灌注48h时肾功能已经有部分恢复。与Sham组比较,缺血再灌注24h肾脏组织肾小管管腔扩张,内有管型形成,肾小管上皮细胞肿胀坏死,空泡变性,刷状缘坏死脱落,并且伴有炎性细胞侵润,而肾小球未见明显病变。免疫组织化学结果显示,与Sham组比较,缺血再灌注24h肾脏VEGF-C及其受体VEGFR-3的表达均明显增加,二者存在着共定位现象,且主要表达在肾脏皮髓质交界处及髓质部的肾小管。Westernblot结果显示随着缺血后再灌注时间的延长,VEGF-C的表达增加。结论肾缺血再灌注损伤后存在肾脏VEGF-C的表达上升,且与其受体VEGFR-3的表达增加呈共定位,推测VEGF-C可能参与了肾脏缺血再灌注损伤。     

Curcumin protects against ischemia/reperfusion injury in rat kidneys

OBJECTIVES: Renal ischemia followed by reperfusion leads to acute renal failure in both native kidneys and renal allograft. We investigated the effect of curcumin on ischemia-reperfusion (I/R) injury and the antioxidant effects of curcumin in rats. METHODS: Thirty rats were randomly divided into five experimental groups (control, sham, curcumin, I/R and I/R+curcumin, n=6 each). Curcumin was administered (200 mg kg(-1)) orally to curcumin and I/R+curcumin groups for 7 days. Then, the rats were subjected to bilateral renal ischemia for 45 min and followed by reperfusion for 24 h. All rats were killed and kidney function tests, serum and tissue nitric oxide (NO), protein carbonyl (PC), malondialdehyde (MDA), superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) levels were determined. Histopathological examinations were also performed. RESULTS: Curcumin significantly improved the urea and cystatin C levels in I/R+curcumin group compared to I/R group (p<0.05). Reduction of serum GSH-Px was significantly improved by curcumin (p<0.001), but SOD enzyme activity did not alter (p>0.05). Treatment with curcumin also resulted in significant reduction in serum and tissue MDA, NO and PC and for tissue that were increased by renal I/R injury (p<0.001 for serum and p<0.05 for tissue, respectively). In histological examination, the rats treated with curcumin had nearly normal morphology of the kidney. CONCLUSIONS: Based on our results, it can be concluded that curcumin protects the kidneys against I/R injury via its antioxidant effects.