热休克蛋白70和血红素加氧酶-1表达在肾缺血后处理减轻肾缺血再灌注损伤中的作用

目的 评价热休克蛋白70(HSP70)和血红素加氧酶-1(HO-1)表达在肾缺血后处理减轻肾缺血再灌注损伤中的作用.方法健康雄性SD大鼠140只,体重250～280 g,采用随机数字表法,将大鼠随机分为4组(n=35):假手术组(S组)仅开腹,游离双侧肾脏,分离双侧肾蒂不夹团;肾缺血再灌注组(I/R组)夹闭双侧肾蒂缺血45 min,恢复灌注;缺血后处理组(IPo组)夹闭双侧肾蒂45 min,再灌注10 s,缺血10 s,反复3次,恢复灌注;HSP抑制剂槲皮黄酮+缺血后处理组(Q+IPo组)缺血前1 h 腹腔注射槲皮黄酮100 mg/kg,余操作同IPo组.于再灌注即刻(T0)、1、3、6、12、24、48 h(T1～6)时各组随机取5只大鼠抽心脏血后取肾,检测肾组织HSP70、HO-1的mRNA和蛋白表达,T3时抽心脏血,测定血清肌酐(Cr)和尿素氮(BUN)浓度、caspase-3 mRNA的表达,TUNNEL法检测肾组织凋亡细胞,计算凋亡指数(AI),光镜下观察肾组织病理学结果.结果 与S组比较,其余组T3时血清Cr和BUN浓度和AJ升高,caspase-3 mRNA表达上调,各时点HSF70、BO-1的mRNA和蛋白表达上调(P＜0.05);与I/R组比较,IPo组T3时血清Cr和BUN浓度和AI降低,caspase-3 mRNA表达下调,T1～5时HSP70、HO-1的mRNA和蛋白表达上调(P＜0.05);与IPo组比较,Q+IPo组T3时血清Cr和BUN浓度和AJ升高,caspase-3mRNA表达上调,T1～5时HSP70、HO-1的mRNA和蛋白表达下调(P＜0.05).IPo组肾组织病理学损伤较I/R组减轻,Q+IPo组肾组织病理学损伤程度与I/R组相似.结论 HSP70和H0-1表达参与了肾缺血后处理减轻肾缺血再灌注损伤的过程.

Abstract：

Objective To evaluate the role of the expression of heat shock protein 70 (HSP70) and heme oxygenase-1 (HO-1) in the reduction of renal ischemia-reperfusion (I/R) injury by ischemic postconditioning in tats.Methods One hundred and forty healthy male SD rats weighing 250-280 g were randomized into 4 groups ( n = 35 each) : sham operation group (S group) ; I/R group; ischemic postconditioning group (IPo group); quercetin (an inhibitor of HSP) + ischemic postconditioning group (Q + IPo group). Renal I/R was produced by clamping bilateral renal pedicels for 45 min followed by reperfusion. In group S, bilateral kidneys were only exposed through a midline incision but their- pedicels were not clamped. In IPo and Q + IPo groups, 45 min ischemia was followed by three 10 s episodes of ischemia at 10 s intervals for reperfusion and in addition intraperitoneal quercetin 100 mg/kg was injected at 1 h before ischemia in group Q + IPo. Blood samples from hearts were obtained at 0, 1, 3, 6, 12, 24 and 48 h of reperfusion (T0-6) and the rats were then sacrificed and kidneys removed to detect the expression of HSP70 and HO-1 mRNA and protein in renal tissues. The blood samples obtained at T3 were used to determine serum creatinine (Cr) and urea nitrogen (BUN) concentrations and the expression of caspase-3 mRNA . The apoptosis in the renal tissues was detected using TUNEL and apoptotic index ( AI) was calculated. Microscopic examination was performed with light microscope. Results Compared with group S, the serum Cr and BUN concentrations and AI were significantly increased at T3,the expression of caspase-3 mRNA was up-regulated at T3, and the expression of HSP70 and HO-1 mRNA and protein was up-regulated at T0-6 in the other groups (P ＜ 0.05) . Compared with group I/R, the serum Cr and BUN concentrations and AI were significantly decreased at T3, the expression of caspase-3 mRNA was down-regulated at T3, and the expression of HSP70 and HO-1 mRNA and protein was up-regulated at T1-5 in group IPo ( P ＜ 0.05) . Compared with group IPo, the serum Cr and BUN concentrations and AI were significantly increased at T3, the expression of caspase-3 mRNA was up-regulated at T3, and the expression of HSP70 and HO-1 mRNA and protein was down-regulated at T1-5, in group Q + IPo ( P ＜ 0.05) . The microscopic examination showed that the renal I/R injury was significantly attenuated by ischemic postconditioning and the degree of injury in group IPo was similar to that in group I/R. Conclusion The expression of HSP70 and HO-1 is involved in the reduction of renal I/R injury by ischemic postconditioning in rats.     

七氟醚延迟预处理对大鼠缺血再灌注心肌ARC表达的影响

目的 评价七氟醚延迟预处理对大鼠缺血再灌注心肌带有Caspase富集功能域的凋亡抑制蛋白(ARC)表达的影响.方法 成年雄性SD大鼠64只,体重270～350 g,采用随机数字表法,将其随机分为4组(n=16):假手术组(S组)、心肌缺血再灌注组(I/R组)、七氟醚+假手术组(S-S组)和七氟醚延迟预处理+心肌缺血再灌注组(S-VR组).S-S组和S-I/R组分别吸入33%氧气和2.5%七氟醚2 h,停止吸入后24 h行假手术或心肌缺血再灌注;I/R组和S-I/R组采用结扎左冠状动脉前降支30 min,再灌注2 h的方法制备心肌缺血再灌注模型.于再灌注2 h时处死8只大鼠,取左心室组织,测定心肌梗死范围及细胞凋亡情况,计算凋亡指数,于缺血前即刻及再灌注2 h时各处死4只大鼠,取左心室组织,测定ARC及Caspase-8的表达水平.结果 与S组比较,I/R组和S-I/R组心肌梗死范围及细胞凋亡指数升高,缺血前即刻S-S组和S-I/R组ARC表达上调,再灌注2 h时I/R组Caspese-8、表达上调(P＜0.05);与I/R组比较,S-I/R组心肌梗死范围和细胞凋亡指数降低,再灌注2 h时S-S组和S-I/R组ARC表达上调,Caspase-8表达下调(P＜0.05).结论 七氟醚延迟预处理可上调心肌ARC表达,减少细胞凋亡的发生,从而减轻大鼠心肌缺血再灌注损伤.

Abstract：

Objective To investigate the effects of sevoflurane delayed preconditioning on caspase recruitment domain (ARC) expression during myocardial ischemia-reperfusion (I/R) in rats. Methods Sixty-four adult male SD rats weighing 270-350 g were randomly divided into 4 groups ( n = 16 each): sham operation (group S); myocardial I/R group; sevoflurane + sham operation group (group S-S) and sevoflurane delayed preconditioning + myocardial I/R group (group S-I/R) . Myocardial I/R was induced by occlusion of anterior descending branch of left coronary artery for 30 min followed by 2 h of reperfusion in groups I/R and S-I/R. Group S-S inhaled 33% oxygen for 2 h, and sham operation was performed 24 h later. Group S-I/R inhaled 2.5% sevoflurane for 2 h, and then myocardial I/R was induced 24 h later. Eight animals were sacrificed at the end of 2 h reperfusion in each group and the hearts removed for determination of myocardial infarct size (IS) as a percentage of area at risk (AAR) by triphenyl tetrazolium chloride staining (IS/AAR) . Myocardial apoptosis was detected using TUNEL and apoptosis index was calculated. Another 4 animals were sacrificed immediately before ischemia and at the end of 2 h reperfusion to determine the expression of ARC and Caspase-8 in myocardium by Western blot. Results Compared with group S, the infarct size and apoptosis index were significantly increased in groups I/R and S-I/R, and ARC expression was up-regulated immediately before ischemia in groups S-S and S-I/R, and Caspase-8 expression was up-regulated at 2 h of reperfusion in group I/R ( P ＜ 0.05) . Compared with group I/R, the infarct size and apoptosis index were significantly decreased in group S-I/R, and ARC expression was up-regulated, while Caspase-8 expression was down-regulated at 2 h of reperfusion in groups S-S and S-I/R ( P ＜ 0.05) . Conclusion Sevoflurane delayed preconditioning can attenuate myocardial I/R injury through up-regulating the ARC expression and decreasing the myocardial apoptosis.     

七氟醚预处理对大鼠肾缺血再灌注损伤的影响

目的 评价七氟醚预处理对大鼠肾缺血再灌注损伤的影响.方法 雄性SD大鼠24只,体重250～300 g,采用随机数字表法,将大鼠随机分为3组(n=8):假手术组(S组)、肾缺血再灌注组(I/R组)和七氟醚预处理组(SP组).I/R组和SP组采用切除右肾然后夹闭左侧肾动脉45 min再开放的方法 制备肾缺血再灌注模型.SP组吸入2.2%七氟醚1 h,停止吸入后10 min时进行肾缺血.于再灌注2 h时采集静脉血样,测定血清肌酐(Cr)、尿素氮(BUN)和胱抑素C(Cys C)的浓度,取肾组织,光镜下及透射电镜下观察病理学结果,并根据肾小管病变程度进行Paller评分.结果 与S组比较,I/R组血清Cr和BUN浓度差异无统计学意义(P＞0.05),血清Cys C浓度和Paller评分明显升高(P＜0.05);与I/R组比较,SP组血清Cys C浓度和Paller评分明显降低(P＜0.05).SP组肾组织损伤程度轻于I/R组.结论 七氟醚预处理可减轻大鼠肾缺血再灌注损伤.

Abstract：

Objective To investigate the effects of sevoflurane preconditioning on renal ischemia-reperfusion(I/R)injury in rats.Methods Twenty-four adult male SD rats weighing 250-300 g were randomly divided into 3 groups(n=8 each):sham operation group (group S);I/R group; sevoflurane preconditioning group (group SP). After the rats underwent right nephrectomy, renal I/R was produced by occlusion of left renal artery for 45 min followed by reperfusion in I/R and SP groups.In group SP, the rats inhaled 2.2% sevoflurane for 1 h, then the inhalation was stopped and renal ischemia was performed 10 min later. Venous blood samples were collected at 2 h of reperfusion to determine the concentrations of serum creatinine(Cr), urea nitrogen (BUN), cystatin C (Cys C) . The renal tissues were obtained for microscopic examination, and Paller's score was recorded. Results Compared with group S, there was no significant difference in the serum Cr and BUN concentrations (P＞0.05), while the serum Cys C concentration and Paller's score for acute renal tubular injury were significantly increased in group I/R(P＜0.05). The serum Cys C concentration and Paller's score were significantly lower in group SP than in group I/R(P＜0.05).I/R-induced renal injury was significantly reduced in group SP compared with group I/R. Conclusion Preconditioning with sevoflurane can provide significant protection against renal I/R injury.     

七氟醚预处理对局灶性脑缺血再灌注损伤大鼠皮质C/EBP同源蛋白表达的影响

目的 探讨七氟醚预处理对局灶性脑缺血再灌注损伤大鼠皮质C/EBP同源蛋白(CHOP)表达的影响.方法 健康雄性SD大鼠36只,体重250～280 g,采用随机数字表法,将大鼠随机分为3组(n=12):假手术组(S组)、局灶性脑缺血再灌注组(I/R组)和七氟醚预处理组(Sevo-pc 组).采用线栓法阻断右侧大脑中动脉1 h,再灌注24 h,制备局灶性脑缺血再灌注损伤模型.Sevo-pc 组于缺血前1 h吸入2.7%七氟醚.各组于再灌注24 h时行神经功能缺陷评分后断头取脑,TIC染色法测定脑梗死体积,免疫组化法测定缺血侧皮质CHOP表达,TUNEL法计数凋亡神经细胞.结果 与S组比较,I/R组和Sevo-pc组神经功能缺陷评分升高,脑梗死体积比升高,缺血侧皮质CHOP表达上调,凋亡细胞数增加(P＜0.01);与I/R组比较,Sevo-pc组神经功能缺陷评分降低,脑梗死体积比降低,缺血侧皮质CHOP表达下调,凋亡细胞数减少(P＜0.05或0.01).结论 七氟醚预处理可能通过下调皮质CHOP表达减轻大鼠局灶性脑缺血再灌注损伤.

Abstract：

Objective To investigate the effect of sevoflurane preconditioning on CCAAT/enhancer binding protein (C/EBP) homologous protein (CHOP) expression in the cerebral cortex after focal cerebral ischemiareperfusion (I/R) injury in rats and the mechanism. Methods Thirty-six male SD rats weighing 250-280 g were randomly divided into 3 groups ( n = 12 each) : sham operation group (group S) , focal cerebral I/R group (group I/R) and sevoflurane preconditioning group (group Sevo-pc). The animals were anesthetized with intraperitoneal chloral hydrate 300 mg/kg. In groups I/R and Sevo-pc, focal cerebral ischemia was induced by middle cerebral artery occlusion using a nylon thread with rounded tip inserted into the right internal carotid artery and advanced cranially until resistance was met. The occlusion was maintained for 1 h followed by 24 h reperfusion. Group Sevo-pc inhaled 2.7% sevoflurane for 1 h before ischemia. Neurological deficits were assessed and scored at the end of 24 h reperfusion and then the rats were decapitated. Their brains were immediately removed. The cerebral infarct size was determined by TTC staining. The CHOP expression in the ischemic cerebral cortex was determined by immunohistochemistry. The number of apoptotic neurons was counted using TUNEL. Results The neurological deficit scores were significantly higher, the cerebral infarct size was significantly larger, and the CHOP expression and the number of apoptotic neurons were significantly higher in groups I/R and Sevo-pc than in group S ( P ＜ 0.01) . The neurological deficit scores were significantly lower, the cerebral infarct size was significantly smaller, and the CHOP expression and the number of apoptosis neurons were significantly lower in group Sevo-pc than in group I/R ( P ＜ 0.05 or 0.01) . Conclusion Sevoflurane preconditioning may protect the brain against focal cerebral I/R injury by down-regulating CHOP expression in the cerebral cortex in rats.     

PI3K-Akt信号通路在七氟醚预处理减轻大鼠离体心脏缺血再灌注损伤中的作用

目的 评价磷脂酰肌醇-3-激酶-丝氨酸/苏氨酸激酶(PI3K-Akt)信号通路在七氟醚预处理减轻大鼠离体心脏缺血再灌注损伤中的作用.方法 健康成年雄性SD大鼠96只,体重220～280g,采用随机数字表法,将其随机分为6组(n=16):假手术组(S组)、缺血再灌注组(I/R组)、七氟醚预处理组(SP组)、渥曼青霉素组(W组)、二甲基亚砜组(D组)和七氟醚预处理+渥曼青霉素组(SW组).采用Langendorff装置建立大鼠离体心脏缺血再灌注模型.S组继续灌注180 min;I/R组平衡灌注30 min,缺血30 min,恢复灌注120 min;其余各组先平衡灌注15 min,SP组、W组、DMSO组和SW组分别用含2.4%七氟醚、100 nmol/L渥曼青霉察、20 μmol/L二甲基亚砜、2.4%七氟醚和100 nmol/L渥曼青霉素的K-H液灌注10 min,然后洗脱5 min,缺血30 min,恢复灌注120 min.各组随机取8个心脏,于平衡灌注末和再灌注15 min时,记录HR、左室舒张末压(LVEDP)、左室发展压(LVDP)、左心室内压最大上升速率(+dp/dtmax)和左心室内压最大下降速率(-dp/dtmax).再灌注15 min时取心肌组织,采用TUNEL法检测细胞凋亡,计算凋亡指数;采用Western blot法测定磷酸化Akt(p-Akt)表达.再灌注120 min时,取8个心脏,采用TIC染色法测定心肌梗死体积.结果 与S组比较,其余各组HR、LVDP和±dp/dtmax降低,LVEDP升高,I/R组、SP组和D组心肌p-Akt表达上调(P＜0.05);与I/R组比较,SP组LVDP和±dp/dtmax升高,LVEDP和凋亡指数降低,心肌p-Akt表达上调,心肌梗死体积减小(P＜0.05),SW组上述指标差异无统计学意义(P＞0.05).结论 七氟醚预处理可通过激活PI3K-Akt信号通路减轻大鼠离体心脏缺血再灌注损伤.

Abstract：

Objective To investigate the role of phosphatidyl-inositol 3-kinase-Akt (PI3k-Akt) signal pathway in the attenuation of ischemia-reperfusion (I/R) injury by sevoflurane preconditioning in isolated rat hearts. Methods Ninety-six adult male SD rats weighing 220-280 g were randomly divided into 6 groups ( n = 16 each): sham operation group (group S); I/R group; sevoflurane preconditioning group (group SP); wortmannin group (group W); dimethyl sulfoxide (DMSO) group (group D) and sevoflurane preconditioning + wortmannin group (group SW) . Their hearts were excised and perfused in a Langendorff apparatus with K-H solution saturated with 95%O2-5%C02 at 37 ℃ . The hearts were continuously perfused for 180 min in group S. After 15 min of equilibration, the isolated hearts were subjected to 30 min of ischemia followed by 120 min of reperfusion in SP, W, D and SW groups. Croups SP, W, D and SW received 10 min of perfusion with K-H solution containing 2. 4% sevoflurane, 100 nmol/L wortmannin, 20 μmol/L DMSO, and 2.4% sevoflurane + 100 nmol/L wortmannin, respectively, followed by 5 min washout before I/R. Eight hearts in each group were selected and HR, left ventricular end-diabetic pressure (LVEDP), left ventricular developed pressure (LVDP), and ± dp/dtmax were recorded at the end of equilibration and at 15 min of reperfusion, Myocardial tissues were obtained at 15 min of reperfusion for determination of apoptosis (by TUNEL) and phosphorylated Akt (p-Akt) expression (by Western blot) . Another 8 hearts were selected at 120 min of reperfusion for determination of myocardial infarct size by TTC staining. Result Compared with group S, LVDP and ± dp/dt,^ were significantly decreased and LVEDP was significantly increased in groups I/R, SP, W, D and SW, and myocardial p-Akt expression was up-regulated in groups I/R, SP and D ( P ＜ 0.05). Compared with group I/R, LVDP and ± dp/dtmax were significantly increased, LVEDP and apoptosis index were significantly decreased, myocardial p-Akt expression was up-regulated, and myocardial infarct size was significantly reduced in group SP (P ＜0.05) . Conclusion Activation of PI3K-Akt signal pathway is involved in the attenuation of I/R injury by sevoflurane reconditioning in isolated rat hearts.     

细胞穿透肽PEP-1介导血红素加氧酶-1对大鼠肠缺血再灌注损伤的影响

目的 评价细胞穿透肽PEP-1介导血红素加氧酶-1(HO-1)对大鼠肠缺血再灌注损伤的影响.方法 雄性SD大鼠18只,周龄7～9周,体重210～260 g,采用随机数字表法,将大鼠随机分为3组(n=6):假手术组(S组)、肠缺血再灌注组(IR组)和融合蛋白PEP-1/HO-1+肠缺血再灌注组(HO组).采用夹闭肠系膜上动脉45 min,恢复灌注120 min的方法制备大鼠肠缺血再灌注损伤模型.HO组夹闭肠系膜上动脉前30 min,左侧髂静脉注射融合蛋白PEP-1/HO-1 0.5 mg,S组不夹闭肠系膜上动脉,余操作同IR组.于再灌注120 min时处死大鼠取小肠组织,称重后计算肠湿/干重比,测定丙二醛(MDA)含量、超氧化物歧化酶(SOD)活性和HO-1活性,免疫组化法检测肠组织HO-1蛋白的表达,光镜下观察肠组织结构并进行损伤评分.结果 与S组比较,IR组和HO组肠湿/干重比和MDA含量升高,SOD活性降低,HO-1活性和蛋白表达水平升高,损伤评分升高(P＜0.05);与IR组比较,HO组肠湿/干重比、MDA含量降低,SOD活性升高,HO-1活性和蛋白表达水平升高,损伤评分降低(P＜0.05).HO组大鼠肠组织病理学损伤较IR组减轻.结论 细胞穿透肽PEP-1可将HO-1成功导人大鼠肠组织中的细胞并减轻肠缺血再灌注损伤.

Abstract：

Objective To investigate the effects of heme oxygenase-1 (HO-1) mediated by cell penetrating peptide PEP-1 on intestinal ischemia/reperfusion (I/R) injuiy in tats. Methods Eighteen male SD rats aged 7-9 weeks weighing 210-260 g were randomly divided into 3 groups (re = 6 each): sham operation group (group S) , I/R group and PEP-1/HO-1 + I/R group (group HO) . To establish a model of intestinal I/R injury, intestines were exteriorized and the superior mesenteric artery was exposed and occluded for 45 min ischemia, and then the clamp was removed for 120 min reperfusion. The PEP-1/HO-1 fusion protein 0.5 mg was injected via the left iliac vein 30 min prior to ischemia in group HO. The superior mesenteric artery was exposed but not occluded in group S. At the end of reperfusion, the rats were sacrificed and intestinal tissues obtained to determine the intestinal wet/ dry ratio, malondialdehyde (MDA) level, activities of superoxide dismutase (SOD) and HO-1, and HO-1 protein expression. The histological changes in the intestinal mucosa were examined and the injuiy was scored. Results Compared with group S, the intestinal wet/dry ratio, MDA level, HO-1 activity, HO-1 protein expression and injury score were significantly increased, while the SOD activity was significantly decreased in groups I/R and HO ( P ＜ 0.05) . Compared with group I/R, the intestinal wet/dry ratio, MDA level and injury score were significantly decreased, while the SOD activity, HO-1 activity and HO-1 protein expression increased in group HO ( P ＜ 0.05) . The pathologic changes were significantly attenuated in group HO compared with group I/R.Conclusion HO-1 protein can be successfully delivered into intestinal tissues by PEP-1 and has protective effects against intestinal I/R injury.     

辛伐他汀预处理对肢体缺血再灌注诱发肺损伤大鼠肺组织血红素加氧酶-1表达的影响

目的 评价辛伐他汀预处理对肢体缺血再灌注诱发肺损伤大鼠肺组织血红素加氧酶-1(HO-1)表达的影响.方法 成年雄性SD大鼠48只,体重250～300 g,采用随机数字表法,将大鼠随机分为6组(n=8):假手术组(S组)、肢体缺血再灌注组(IR组)、辛伐他汀1、5、10 mg/kg组(S1组、S2组、S3组)和辛伐他汀对照组(SC组).采用夹闭股动脉2 h,再灌注3 h的方法制备肢体缺血再灌注模型.S组:仅分离股动脉和股静脉,不夹闭;IR组:制备肢体缺血再灌注模型;S1组、S2组、S3组:分别将辛伐他汀1、5、10 mg/kg溶于1 ml蒸馏水,于每13清晨灌胃1次,连续灌胃3 d后制备肢体缺血再灌注模型;SC组:辛伐他汀10 mg/kg溶于1 ml蒸馏水,于每日清晨灌胃1次,连续灌胃3 d.再灌注3 h时取颈动脉血样,行血气分析,记录PaO2和PaCO2,随后处死大鼠,取肺组织,观察病理学结果,计算湿重/干重比(W/D比),测定SOD活性,计数PMN,测定HO-1 mRNA及其蛋白的表达水平.结果 与S组比较,IR组PaO2及PaCO2降低,IR组、S1组和S2组肺组织W/D比和PMN计数升高,SOD活性降低(P＜0.05),S3组和SC组上述指标差异无统计学意义(P＞0.05),IR组、S1组、S2组、S3组和SC 组肺组织H0-1 mRNA及其蛋白表达上调(P＜0.01);与IR组比较,S1组、S2组和S3组PaO2、PaCO2及肺组织SOD活性升高,肺组织W/D比和PMN计数降低,肺组织HO-1 mRNA及其蛋白表达上调(P＜0.05或0.01);S1组、S2组和S3组肺组织WID比和PMN计数依次降低,SOD活性依次升高,HO-1 mRNA及其蛋白表达依次上调(P＜0.05或0.01).S1组、S2组和S3组肺组织病理性损伤较IR组减轻.结论 辛伐他汀预处理可上调肢体缺血再灌注大鼠肺组织HO-1的表达,从而产生肺保护作用,且呈剂量依赖性.

Abstract：

Objective To investigate the effects of simvastatin preconditioning on the pulmonary heme oxygenase-1 (HO-1) expression in rats with lung injury induced by ischemia-reperfusion (I/R) of hind limbs. Methods Forty-eight adult male SD rats weighing 250-300 g were randomly divided into 6 groups ( n = 8 each) : sham operation group (group S) ; I/R group; I/R + simvastatin 1,5, 10 mg/kg groups (S1 , S2, S3 groups) ; simvastatin control group (group SC) . I/R of hind limbs was produced by occlusion of bilateral femoral arteries for 2 h followed by 3 h reperfusion. Croups S1 , S2 , S3 received simvastatin 1, 5, 10 mg/kg respectively via an oro-gastric tube for 3 days before I/R. Group SC received simvastatin 10 mg/kg via an oro-gastric tube for 3 days. Arterial blood samples were taken at 3 h of reperfusion for blood gas analysis and PaO2 and PaCO2 were recorded. The animals were then sacrificed and the lungs removed immediately for pathologic examination and determination of the wet/dry lung weight ratio (W/D ratio), superoxide dismutase (SOD) activity and polymorphonuclear neutrophil (PMN) count . Hie expression of HO-1 mRNA and protein in lung tissues was detected using RT-PCR and Western blot analysis respectively.Results Alveolar edema, localized pulmonary atelectasis and large amount of PMN infiltration were found in I/R group and were ameliorated in S1, S2, S3 groups. Compared with group S, PaO2 and PaCO2 were significantly decreased in I/R group, W/D ratio and PMN count were increased and SOD activity was significantly decreased in I/R, S1 , S2 groups, and expression of HO-1 mRNA and protein was up-regulated in the other five groups ( P ＜ 0.05). PaO2, PaCO2 and SOD activity were significantly increased, W/D ratio and PMN count were significantly decreased, and HO-1 mRNA and protein expression was up-regulated in S1, S2 and S3 groups as compared with I/R group ( P ＜ 0.05 or 0.01). W/D ratio and PMN count were gradually decreased, SOD activity was gradually increased, and HO-1 mRNA and protein expression was gradually up-regulated in S1, S2 and S3 groups. Conclusion Simvastatin preconditioning has protective effect against lung injury induced by I/R of hind limbs in rats through up-regulation of HO-1 expression in the lung tissues and in a dose-dependent manner.     

七氟醚麻醉对新生大鼠杏仁核DNA甲基转移酶mRNA表达的影响

目的 探讨七氟醚麻醉对新生大鼠杏仁核DNA甲基转移酶(DNMT)mRNA表达的影响.方法 新生(出生8 d)SD大鼠42只,采用随机数字表法,将其随机分为2组(n=21),对照组:不给予任何处理;实验组:吸入5%七氟醚1 min后,吸入浓度调整为3%维持4 h.实验组分别于停止吸入即刻及停止吸入后24 h时,处死大鼠,取杏仁核,测定DNMT1 mRNA、DNMT3a mRNA、DNMT3b mRNA的表达;停止吸入即刻取动脉血样,进行血气分析.对照组于相应时间点进行处理.结果 与对照组比较,实验组DNMT1 mRNA、DNMT3a mRNA表达均下调(P＜0.05),DNMT3b mRNA表达和血气分析指标差异无统计学意义(P＞0.05).结论 七氟醚麻醉可下调新生大鼠杏仁核DNMT1 raRNA和DNMT3a mRNA的表达,该作用可能会导致发育期中枢神经功能损伤.

Abstract：

Objective To investigate the effect of sevoflurane anesthesia on DNA methyltransferase (DNMT) mRNA expression in neonatal rat amygdala. Methods Forty-two 8-day-old SD rats were randomly divided into 2 groups ( n = 21 each): control group and experimental group. 5% sevoflurane was inhaled for 1 min, and then the inhaled concentration of sevoflurane was decreased to 3 % and maintained for 4 h. The rats were sacrificed at the end of sevoflurane inhalation and 24 h after the end of sevoflurane inhalation, and amygdala was removed for determination of DNMT, mRNA, DNMT3, mRNA and DNMT3b mRNA expression by RT-PCR. Blood samples were taken at the end of sevoflurane inhalation for blood gas analysis. Results Compared with control group, the DNMT, mRNA and DNMT3, mRNA expression was down-regulated (P ＜ 0.05). There were no significant differences in DNMT3b mRNA expression and parameters of blood gas analysis between the two groups (P ＞ 0.05). Conclusion Sevoflurane anesthesia can down-regulate DNMT, mRNA and DNMT,, mRNA expression in neonatal rat amygdala, which may result in functional deficits during the development of central nervous system.     

不同浓度七氟醚预处理对大鼠海马神经元缺氧复氧时细胞凋亡的影响及线粒体ATP敏感型钾通道在其中的作用

目的 探讨不同浓度七氟醚预处理对大鼠海马神经元缺氧复氧时细胞凋亡的影响及线粒体ATP敏感型钾通道(mito-KATP通道)在其中的作用.方法 新生(出生＜24 h)SD大鼠,雌雄不拘,体重5～6 g,原代培养海马神经元,接种于培养孔或培养皿中,采用随机数字表法,将其随机分为7组,每组48孔和12皿,正常对照组(C组):不予任何处理;缺氧复氧组(HR组):缺氧4 h复氧24 h;6%七氟醚预处理组(S1 组)、4%七氟醚预处理组(S2 组)、2%七氟醚预处理组(S3 组):分别经6%、4%、2%七氟醚预处理后行缺氧复氧;5-羟葵酸100 μmol/L预处理组(5-HD组):经mito-KATP通道阻断剂5-羟葵酸(终浓度100 μmol/L)预处理后进行缺氧复氧;5-羟葵酸100 μmol/L+6%七氟醚预处理组(5-HD+S组):同时行5-羟葵酸和6%七氟醚预处理后进行缺氧复氧.各组以上处理结束后,测定神经元活力、凋亡率、Bcl-2和Bax蛋白的表达水平.结果 与C组比较,其余6组海马神经元活力降低,细胞凋亡率升高,Bcl-2和Bax蛋白表达上调(P＜0.01);与HR组比较,S1组～S3组海马神经元活力增强,细胞凋亡率降低,Bcl-2蛋白表达上调,Bax蛋白表达下调(P＜0.01),5-HD组和5-HD+S组上述指标比较差异无统计学意义(P＞0.05);与S1组比较,S2组、S3组和5-HD+S组海马神经元活力降低,细胞凋亡率升高,Bcl-2蛋白表达下调,Bax蛋白表达上调(P＜0.01);与S2组比较,S3组海马神经元活力降低,细胞凋亡率升高,Bcl-2蛋白表达下调,Bax蛋白表达上调(P＜0.01).结论 七氟醚预处理可抑制大鼠海马神经元缺氧复氧时细胞凋亡,从而减轻神经元损伤,且呈浓度依赖性,机制可能与开放神经元mito-KATP通道,上调Bcl-2蛋白表达,下调Bax蛋白表达有关.

Abstract：

Objective To investigate the effect of preconditioning with different concentrations of sevoflurane on hypoxia-reoxygenation(H/R)-induced apoptosis in rat hippocampal neurons and the role of mitochondrial KATP(mito-KATP)channels.Methods Primary cultured hippocampal neurons isolated from newborn SD rats(＜24h)of both sexes,weighing 5-6 g,were randomly divided into 7 groups with 48 wells and 12 dishes in each one:control group(C group),H/R group,preconditioning with 6%,4%and 2% sevoflurane groups(S1-3 groups),5-hydroxydecanoate(5-HD,mito-KATP channel blocker)100 μmol/L preconditioning group(5-HD group)and preconditioning with 5-HD 100 μmol/L+6% sevoflurane group(5-HD+S group).The neurons were exposed to 4 h hypoxia followed by 24 h reoxygenation. In S1-3 groups, preconditioning was performed with 6% , 4% and 2% sevoflurane respectively before H/R. In 5-HD group, preconditioning was performed with 5-HD (final concentration 100 μmol/L) before H/R. In 5-HD + S group, preconditioning was performed with 5-HD 100 μmol/L and 6% sevoflurane before H/R. The neuronal viability, apoptosis rate and expression of Bcl-2 and Bax were determined after 24 h reoxygenation.Results The neuronal viability was significantly lower,while the apoptosis rate and expression of Bcl-2 and Bax were significantly higher in the other 6 groups than in group C(P＜0.01).The neuronal viability and expression of Bcl-2 were significantly higher,while the apoptosis rate and Bax expression were lower in S1-3 groups than in group H/R. There was no significant difference in the parameters mentioned above between 5-HD and 5-HD + S groups(P＞0.05).The neuronal viability and expression of Bcl-2 were significantly lower, while the apoptosis rate and Bax expression were higher in S2, S3 and 5-HD + S groups than in group S1, and in group S3 than in group S2(P＜0.0l) .Conclusion Sevoflurane preconditioning can inhibit H/R-induced apoptosis in rat hippocampal neurons and reduce the injury to neurons in a concentration-dependent manner, and the underlying mechanism may be related to activation of mito-KATP channels, up-regulation of Bcl-2 expression and down-regulation of Bax expression.     

七氟醚预处理联合后处理对大鼠心肌缺血再灌注时血栓素A2和前列腺素I2的影响

Objective To investigate the effect of sevoflurane preconditioning-postconditioning on thromboxane A2 and prostaglandin I2 during myocardial ischemia-reperfusion (I/R) in rats. Methods Fifty healthy male Wistar rats weighing 250-280 g were randomly divided into 5 groups (n = 10 each) : sham operation group (group S) , I/R group, sevoflurane preconditioning group (group Spr), sevoflurane postconditioning group (group Spo)and combination of sevoflurane preconditioning and postconditioning group (group Spr + po). Myocardial I/R was produced by occlusion of anterior descending branch of left coronary artery for 30 min followed by 2 h reperfusion in anesthetized rats. In group S the anterior descending branch was only exposed but not ligated. Group Spr received 15 min inhalation of 2.5 % sevoflurane and 15 min wash-out 30 min before ischemia. Group Spo received 5 min inhalation of 2.5% sevoflurane 1 min before reperfusion. Arterial blood samples were taken at 2 h of reperfusion for determination of the levels of MB isoenzyme of creatine kinase (CK-MB) , lactate dehydrogenase (LDH) , cardiac troponin I (cTnI), thromboxane B2(TXB2), and 6-keto-prostaglandin (6-keto-PGF1α) and platelet maximum aggregation rate. TXB2/6-keto-PGF1α ratio was calculated. The myocardial tissues were taken for microscopic examination. Mitochondria] injury was assessed by using Flameng score and stereology (Specific surface, δ and Numerical density on area, NA) .Results Compared with group S, the levels of CK-MB, LDH, cTnI, TXE2 and 6-ketoPGF1α, TXB2/6-keto-PGF1α ratio, platelet maximum aggregation rate and Flameng score were significantly increased, while δ and NA were significantly decreased in group I/R (P < 0.05 or 0.01) . The levels of CK-MB,LDH and cTnI, TXB2/6-keto-PGF1α ratio and Flameng score were significantly lower, and 6-keto-PGF1α level, δand NA were significantly higher in Spr and Spo groups than in group I/R ( P < 0.05 or 0.01) . The levels of CKMB, LDH, cTnI and TXB2 , TXB2/6-keto-PGF1α ratio, platelet maximum aggregation rate and Flameng score were significantly lower and 6-keto-PGF1α level,δ and NA were significantly higher in group Spr + po than in Spr and Spo groups(P < 0.05). Conclusion Sevoflurane preconditioning-postconditioning can reduce myocardial I/R injury through inhibiting the release of thromboxane A2 and promoting the release of prostaglandin I2 in rats.     

七氟醚对大鼠急性肾缺血-再灌注损伤的保护作用

目的探讨七氟醚对急性肾缺血-再灌注损伤的保护作用及其机制。方法SD大鼠18只,随机均分为缺血-再灌注组(I/R组)、七氟醚组(S组)和对照组(C组)。建立大鼠急性肾缺血-再灌注损伤模型,缺血-再灌注后3h分别检测血清尿素氮(BUN)、肌酐(Cr)、超氧化物歧化酶(SOD)、丙二醛(MDA)及观察肾组织的病理学变化。结果与C组比较,I/R组和S组血清BUN、Cr水平显著增加(P<0.05),但S组BUN、Cr低于I/R组(P<0.05)。与I/R组比较,S组SOD显著升高,MDA显著降低(P<0.05)。S组肾组织病理损伤分级明显低于I/R组(P<0.05)。结论七氟醚对大鼠急性肾缺血-再灌注损伤具有保护作用,抑制氧自由基反应可能是其重要机制。     

血红素氧合酶-1在感染性休克大鼠肾功能损伤中的作用

目的 探讨血红素氧合酶-1(HO-1)在感染性休克大鼠肾功能损伤中的作用.方法 健康清洁级SD大鼠80只,月龄3～4月,体重260～330 g,雌雄不拘,随机分为4组(n=20):对照组(C组)、感染性休克组(S组)、感染性休克+ZnPP-Ⅸ组(SZ组)和ZnPP-Ⅸ组(Z组).C组静脉注射生理盐水0.5 ml,2 h后腹腔注射50 mmol/L碳酸氢钠溶液1 ml;S组静脉注射LPS 10 mg/kg,2 h后腹腔注射50 mmol/L碳酸氢钠溶液1 ml;SZ组静脉注射LPS 10 mg/kg,2 h后腹腔注射ZnPP-Ⅸ10 μmol/kg;Z组静脉注射生理盐水0.5 ml,2 h后腹腔注射ZnPP-Ⅸ10 μmol/kg.于腹腔注射碳酸氢钠溶液或ZnPP-Ⅸ后4 h时,检测碳氧血红蛋白浓度(COHb)、血清尿素氮(BUN)、肌酐(Cr)的浓度和尿α1-微球蛋白(α1-M)浓度、N-乙酰-a-D-氨基葡萄糖酐酶(NAG)活性、视黄醇结合蛋白(RBP)浓度、γ-谷氨酰转肽酶(γ-GT)活性、肾组织丙二醛(MDA)含量、超氧化物歧化酶(SOD)活性、HO-1 mRNA、HO-2 mRNA、HO-1和HO-2的表达水平.结果 与C组比较,S组和SZ组血清Cr、BUN的浓度、尿α1-M、RBP、NAG、γ-GT水平、COHb、肾组织MDA、HO-1 mRNA、HO-1水平升高,肾组织SOD活性降低(P<0.05),Z组上述指标差异无统计学意义(P>0.05);与S组比较,SZ组血清Cr、BUN的浓度、尿α1-M、RBP、NAG、γ-GT水平和肾组织MDA含量升高,肾组织SOD活性、COHb和肾组织HO-1 mRNA、HO-1水平降低(P<0.05);4组大鼠肾组织HO-2 mRNA、HO-2表达差异无统计学意义(P>0.05).结论 肾组织HO-1对感染性休克大鼠的肾功能可产生一定的保护作用.     

七氟醚麻醉对局灶性脑缺血再灌注大鼠脑组织水通道蛋白9表达的影响

目的 评价七氟醚麻醉对局灶性脑缺血再灌注大鼠脑组织水通道蛋白9( AQP-9)表达的影响.方法 成年雄性SD大鼠75只,体重230～270 g,采用随机数字表法,将其随机分为3组(n=25):假手术组(S组)、缺血再灌注组(UR组)和七氟醚麻醉组(SE组).S组和I/R组静脉注射芬太尼10μg/kg后静脉输注芬太尼25μg·kg-1·h-1,吸入65％N2O和35％O2;SE组吸入2.0％七氟醚和35％ O2.I/R组和SE组采用线栓法制备大鼠局灶性脑缺血再灌注模型,缺血2h后恢复灌注.分别于再灌注6h、1、2、3和5d时测定神经功能缺陷评分和脑组织AQP-9表达,计算脑肿胀率.结果 与S组比较,I/R组和SE组再灌注6h、1、2、3和5d时神经功能缺陷评分升高,再灌注1、2、3d时脑肿胀率升高,再灌注1、2、3和5d时脑组织AQP-9表达上调(P＜0.05);与I/R组比较,SE组再灌注2和3d时神经功能缺陷评分下降,脑肿胀率降低,再灌注2、3和5d时脑组织AQP-9表达上调(P＜0.05).结论 七氟醚麻醉减轻大鼠局灶性脑缺血再灌注损伤的机制与上调脑组织AQP-9的表达有关.     

乌司他丁对重症急性胰腺炎大鼠肾脏细胞凋亡及bcl-2基因表达的影响

目的探讨乌司他丁(UTI)对重症急性胰腺炎(SAP)大鼠肾脏细胞凋亡及bcl-2基因表达的影响。方法60只大鼠随机均分为假手术组、SAP组和UTI组,大鼠SAP模型采用5%牛磺胆酸钠逆行胆胰管注射法建立。于建模后6h及14h时测定血Cr及BUN,于光镜及电镜下观察肾组织病理变化,TUNEL法检测肾脏细胞凋亡情况,SABC免疫组化染色法测定肾脏组织bcl-2基因的蛋白表达。结果SAP组光、电镜下见肾脏组织损害明显,血Cr、BUN、肾脏细胞凋亡指数和bcl-2表达均高于假手术组(P<0.05,P<0.01),且除bcl-2表达外,均随病程延长逐渐升高(P<0.05);经UTI治疗后,光、电镜下见肾脏组织损害减轻,血Cr、BUN和肾脏细胞凋亡指数较SAP组明显下降(P<0.05),而bcl-2表达增强(P<0.05)。肾脏细胞凋亡指数与血BUN和Cr均呈正相关(r=0.807,P<0.05;r=0.812,P<0.05)。结论UTI对肾功能的保护作用可能部分是通过调节凋亡相关基因bcl-2的表达从而影响细胞凋亡实现的。     

Lysophosphatidic acid and lovastatin might protect kidney in renal I/R injury by downregulating MCP-1 in rat

Abstract Renal ischemia/reperfusion (I/R) injury is a major cause of renal failure. The aim of our study is to explore the role of lysophosphatidic acid (LPA) and lovastatin on renal I/R injury and its mechanism in the rat. Male Wistar rats were randomly divided into sham-operated group; renal I/R for 0 h, 4 h, 12 h, and 24 h groups; LPA treatment group; and lovastatin treatment group (n = 10). Rats were killed to determine the level of monocyte chemotactic protein-1 (MCP-1) in renal tissue, renal function [serum creatinine (Cr) and blood urea nitrogen (BUN)], and renal histomorphology to evaluate the effectiveness of LPA and lovastatin. Normal renal tissue had a low level of MCP-1. The level of MCP-1 began to rise at 0 h after reperfusion, reached peak value at 4 h, and then gradually fell off. Compared with sham-operated group, MCP-1 was increased in all renal I/R injury groups (p < 0.01). With the extension of reperfusion, Cr and BUN were significantly increased (p < 0.01). There were damages in kidney tubules, renal interstitium, and kidney glomerulus in renal I/R injury groups. Paller's score was significantly increased in all renal I/R injury groups compared with sham-operated group (p < 0.01). LPA and lovastatin reduced the level of MCP-1, Cr, BUN, and damages of renal histomorphology (p < 0.01). The level of MCP-1 in renal tissue dynamically increases in renal I/R injury, indicating that MCP-1 is involved in renal I/R injury. LPA and lovastatin might protect renal function by downregulating MCP-1 in renal I/R injury.     

血红素加氧酶-1在七氟醚预处理减轻乳鼠心肌细胞缺氧复氧损伤中的作用

目的 探讨血红素加氧酶-1(HO-1)在七氟醚预处理减轻乳鼠心肌细胞缺氧复氧损伤中的作用.方法 新生健康清洁级SD大鼠15只,日龄1～3d,处死后取心室肌组织,原代培养心肌细胞,以1×106个/ml接种于6孔培养板或以2× 105个/ml接种于24孔培养板,采用随机数字表法,将其随机分为4组(n＝25):对照组(C组)常规培养;缺氧复氧组(H/R组)采用缺氧2h,复氧1h的方法制备心肌细胞缺氧复氧损伤模型;七氟醚预处理组(S+ H/R组)细胞经2.5％七氟醚预处理20min后行药物洗脱10 min,再行缺氧复氧处理;锌原卟啉+七氟醚预处理组(ZnPP+ S+ H/R组)细胞经HO-1抑制剂锌原卟啉3 μmol/L孵育1h后,行七氟醚预处理及缺氧复氧处理.于复氧结束后测定心肌细胞HO-1表达、细胞凋亡率、细胞内游离Ca2+浓度([Ca2+]i)、线粒体膜通透性转运孔(PTP)开放程度、细胞色素C(Cyto C)表达及培养液LDH和CK活性.结果 与C组比较,H/R组心肌细胞HO-1和胞浆Cyto C表达上调,线粒体Cyto C表达下调,培养液LDH、CK活性、细胞凋亡率、[Ca2+]i和PTP开放度升高(P<0.01).与H/R组比较,S+H/R组心肌细胞HO-1和线粒体Cyto C表达上调,胞浆Cyto C 表达下调,培养液LDH、CK活性、细胞凋亡率、[Ca2+]i和PIP开放度降低(P<0.01).与S+H/R组比较,ZnPP+ S+ H/R组心肌细胞HO-1和线粒体Cyto C表达下调,胞浆CytoC表达上调,培养液LDH、CK活性、细胞凋亡率、[Ca2+]i和PTP开放度升高(P<0.01).结论 HO-1表达上调参与了七氟醚预处理减轻乳鼠心肌细胞缺氧复氧损伤.     

七氟烷预处理对局灶性脑缺血再灌注损伤大鼠线粒体通透性转换孔的影响

目的 探讨七氟烷预处理对局灶性脑缺血再灌注损伤大鼠线粒体通透性转换孔(mPTP)的影响.方法 成年雄性SD大鼠60只,体重250～300 g,随机分为5组(n=12):假手术组(S组)、缺血再灌注组(I/R组)、七氟烷预处理组(Sev组)、线粒体ATP敏感性钾离子通道(mito-KATP通道)阻断剂5-羟癸酸(5-HD)+Sev组和5-HD组.采用大脑中动脉阻断法制备局灶性脑缺血再灌注模型.S组只分离血管不置入线栓;I/R组制备局灶性脑缺血再灌注模型;Sev组吸入2.4%七氟烷60 min行预处理,24 h后制备局灶性脑缺血再灌注模型;5-HD+Sev组腹腔注射5-HD 40mg/kg,30 min后行七氟醚预处理,其余处理同Sev组;5-HD组腹腔注射5-HD 40 mg/kg,30 min后制备局灶性脑缺血再灌注模型.于再灌注24 h时断头取缺血侧顶叶皮层组织,测定mPTP活性,Western blot法测定Bcl-2、Bax表达水平,并计算Bcl-2/Bax比值,采用TUNEL法检测神经元凋亡情况.结果 与S组比较,I/R组、Sev组、5-HD+Sev组和5-HD组凋亡神经元计数升高,Bcl-2和Bax表达上调,Bcl-2/Bax比值升高,mPTP活性升高(P＜0.05);与I/R组比较,Sev组凋亡神经元计数减少,Bcl-2表达上调,Bcl-2/Bax比值升高,mPTP活性降低(P＜0.05);与Sev组比较,5-HD+Sev组和5-HD组Bcl-2表达下凋,Bcl-2/Bax比值降低,mPTP活性升高(P＜0.05);5-HD+Sev组与5-HD组上述指标比较差异无统计学意义(P＞0.05).结论 七氟烷预处理可能通过激活神经元mito-KATP通道,上调Bcl-2的表达,从而抑制mPTP的大量开放减轻大鼠局灶性脑缺血再灌注时的神经元凋亡.