Calcium-accentuated ischemic damage during reperfusion: the time course of the reperfusion injury in the isolated working rat heart model

The purpose of this study was to determine the time course of calcium-induced postischemic reperfusion injury to the myocardium, using an initial short-term calcium-enriched reperfusion solution. The isolated rat heart model was subjected to 30 min of normothermic potassium cardioplegia-induced ischemic arrest. Control hearts received normal calcium Krebs-Henseliet buffer (KHB) reperfusion. Experimental hearts were challenged with 10 min of calcium-enriched (KHB) reperfusion starting at 0, 1, 2, 5, 15, and 30 min after the beginning of reperfusion. Aortic flow recovery 60 min after reperfusion was used to determine functional recovery. Control hearts recovered 82 +/- 3% of preischemic aortic flow. Hearts which received calcium challenge at 0 and 1 min after the start of reperfusion recovered 43 +/- 4 and 69 +/- 3% of preischemic aortic flow, respectively (P less than 0.01 and P less than 0.05, respectively). Hearts which received calcium challenge 2, 5, 15, and 30 min after reperfusion recovered 75 +/- 2, 80 +/- 2, 85 +/- 2, and 83 +/- 2% of preischemic aortic flow, respectively. Our results indicate that the postischemic myocardium is very susceptible to calcium-accentuated ischemic damage during the initial period of reperfusion. The postischemic heart, however, quickly recovers its ability to withstand a calcium challenge. Five minutes after the start of reperfusion the heart is not influenced by calcium challenge.     

Preoperative bombesin administration can protect the rat small bowel allograft from ischemic reperfusion injury

Background/Purpose

Ischemic reperfusion injury (IR/I) should be minimum for the success of small bowel transplantation (SBTx). This study investigated whether preoperative administration of neuropeptide bombesin (BBS) had a protective effect against IR/I and subsequent acute rejection.

Methods

Allogenic SBTx was performed heterotopically in rats (n = 18). All rats were administered FK506 (0.32 mg/kg per day) everyday. The rats were divided into 3 groups of 6 rats each: group 1, BBS(−)5: warm ischemic time (WIT), 5 minutes without BBS; group 2, BBS(−)15: WIT, 15 minutes without BBS; group 3, BBS(+)15: WIT, 15 minutes with BBS. The specimens were obtained from the stoma site at 1 hour after reperfusion and on postoperative day (POD) 1 and 7. The graft mucosal state and degree of acute rejection were evaluated by H&E staining. The apoptotic cells in the crypt lesion was evaluated using terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling immunohistochemistry. Apoptotic index (AI) was calculated for quantitative analysis.

Results

H&E staining revealed that the mucosal villi on POD 1 remained shortened in the BBS(−)15 group than in the other two groups. One hour after reperfusion, the AI in the BBS(−)15 group was 145.0‰ ± 37.2‰, which was significantly higher (P < .05) than in the BBS(−)5 group (32.6‰ ± 5.0‰) or the BBS(+)15 group (32.0‰ ± 3.0‰). On POD 7, the AI in the BBS(−)15 group was 63.7‰ ± 5.03‰, which was significantly higher (P < .05) than in the BBS(−)5 (17.3‰ ± 4.6‰) or the BBS(+)15 group (12.3‰ ± 3.06‰).

Conclusions

Even a short WIT of 15 minutes induced considerable allograft mucosal damage, which also worsened acute rejection. Exogenous BBS could prevent mucosal damage by IR/I and was also beneficial for the prevention of acute rejection.     

Optimal timing of extracorporeal shock wave treatment to protect ischemic tissue

Enhancement of flap survival through extracorporeal shock wave treatment (ESWT) is a promising new technique; however, no attempt has been made to define the optimal time point and frequency of ESWT to optimize treatment with ESWT for ischemic indications. Twenty-eight male Wistar rats were randomized into 4 groups and an oversized, random-pattern flap was raised and reattached in place in each animal. ESWT was applied 7 days before (group E7) or immediately after the surgical intervention (group E0). The third group was treated with ESWT 7 days before and additionally immediately after the operation (group E7/0). The fourth group served as a control group and did not receive any ESWT (group C). Seven days after flap harvest the results of flap survival, perfusion, microvessel density, and vascular endothelial growth factor concentrations were assessed. Flap survival was significantly increased in all ESWT groups as compared with the control group. The groups (E7 and E0) that received ESWT pre- or postoperatively showed a significant increase in flap perfusion and microvessel density. Combined pre- and postoperative ESWT application (group E0/E7) did not demonstrate a cumulative effect in any evaluation. In this study, we were be able to prove the effectiveness of ESWT in the protection of ischemic tissue flaps. This study suggests that single postoperative application is the most efficacious protocol for clinical applications of ESWT in the treatment of ischemic tissue.     

心麻液预处理的心肌保护效果

目的 研究Ｓｔ．Ｔｈｏｍａｓ晶体心麻液预处理和缺血预处理对大鼠缺血／再灌注心肌收缩功能的影响。方法 应用离体Ｌａｎｇｅｎｄｏｒｆｆ逆行灌注模型,观察晶体心麻液预处理的缺血预处理时心肌缺血及再灌注前后ＬＶＥＤＰ,ＤＰ,＋ｄｐ／ｄｔ－,ＣＦ冠脉流出液中蛋白含量和ＬＤＨ活性以及再灌注后心肌ＳＯＤ活性,ＭＤＡ和ＡＴＰ含量的变化。结果 在缺血期间,晶体心麻液预处理显著延迟了心肌缺血性挛缩的发生时间。     

Protection of acutely ischemic myocardium by controlled reperfusion

The goal of revascularization after acute occlusion of a coronary artery is the return of contractile function and the reduction of mortality. Although reperfusion of ischemic myocardium is a prerequisite for return of function, it may, in itself, cause further injury. Controlled blood cardioplegic reperfusion reduces this "reperfusion injury" and provides maximal myocardial protection. In this article, we review recent advances in surgically controlled reperfusion and speculate on future prospects for myocardial protective techniques in patients with acute coronary artery occlusion.     

Prevention of ischemic damage using controlled limb reperfusion

Following prolonged limb ischemia, a reperfusion injury may occur with the reintroduction of unmodified blood, resulting in tissue loss and, in severe cases, limb loss. We have shown that the reperfusion injury in the heart can be minimized by using controlled reperfusion with a substrate-enriched cardioplegia solution prior to restoring normal blood flow. This article describes two clinical cases in which we used controlled reperfusion in an ischemic limb to prevent limb loss. It demonstrates that a controlled, substrate-enhanced, hypocalcemic, leukodepleted, modified blood reperfusate solution can minimize limb reperfusion damage and improve functional recovery. This preliminary experience is presented to familiarize surgeons with this form of treatment and to describe the solutions and method of administration that can be used to avoid the devastating complications of severe limb ischemia.     

A comparison of ischemic preconditioning versus terminal warm cardioplegia with controlled reperfusion in open heart operation

BACKGROUND: The purpose of this study was to evaluate the effects of three different methods of cardioprotection in patients undergoing valve replacement. METHODS: Ninety patients undergoing elective valve replacement were randomly divided into three groups. In group 1 (n=30), the patients received intermittent cold blood cardioplegia. In group 2 (n=30) they received terminal warm cardioplegia and controlled reperfusion, and in group 3 (n=30), the patients received two cycles of ischemia (2 minutes) and reperfusion (3 minutes) before heart arrest induced by cold blood cardioplegia. The parameters of cardiac function, creatine kinase MB, and clinical outcomes were recorded to assess the effects of experiment. RESULTS: The major preoperative and intraoperative variables are comparable within the three groups. The number of patients requiring the support of inotropic agents was 70% (21/30), 33% (11/30) and 40% (12/30) in group 1, 2 and 3, respectively (p<0.05). The doses of inotropic agent in groups 2 and 3, were significantly lower than in group 1 (1.5+/-0.3 and 1.8+/-0.4 versus 4.5+/-0.8 microg x kg x min(-1), p<0.01) during the first 24 hours after operation. Two deaths (30 day-hospital mortality) occurred, one in group 1 and one in group 2. The cardiac index at 2 hours after bypass discontinuing were 2.2+/-0.04, 3.0+/-0.1 and 2.8+/-0.05 L/m(2) in group 1, 2 and 3, respectively (p<0.01). The left ventricular stroke work index were 24.8+/-1.3, 34.5+/-1.6 and 31.6+/-1.2 g/m x m(2) in group 1, 2, 3, respectively (p<0.01). The release of CK-MB in group 2 and 3 were lower than in group 1 (68+/-7, 81+/-9 versus 116+/-10 IU/L, p<0.01). CONCLUSIONS: Terminal warm cardioplegia with controlled aortic root reperfusion and ischemic preconditioning equally improve cardiac function and reduce the requirement of inotropic agents in patients undergoing valve replacement.     

Can ischemic preconditioning alone really protect organs from ischemia reperfusion injury in transplantation

Organ transplantation is the only choice for treatment of end-stage disease. The ischemia reperfusion injury (I/RI) occurring after cold ischemia is an unavoidable injury during transplantation, which is also one of the main causes of graft failure. Multiple mechanisms have been postulated to explain tissue injury that occurs after I/RI. It is well-known that ischemic preconditioning (IPC), a short period of ischemia followed by reperfusion, arouses the endogenous mechanism of protection against a sustained ischemic insult. Can ischemic preconditioning alone really protect organs from ischemia reperfusion injury in transplantation?     

缺血后处理对大鼠骨骼肌缺血再灌注损伤的影响

目的 探讨缺血后处理对大鼠骨骼肌缺血再灌注损伤的影响以及应用缺血后处理的时机.方法 将32只大鼠随机分成四组,采用切断患肢全部皮肤、肌肉和神经,保留患肢股动静脉的动物模型,通过夹闭和开放股动静脉造成骨骼肌缺血和再灌注损伤.采用测定骨骼肌缺血4 h.再灌注1 h后血清丙二醛(MDA)、骨骼肌髓过氧化物酶(MPO),再灌注6 h后骨骼肌的死亡程度来观察缺血后处理对大鼠骨骼肌缺血再灌注损伤的影响,以及再灌注5 min后应用缺血后处理是否对骨骼肌缺血再灌注损伤有保护作用.结果 对骨骼肌缺血4 h再灌注6 h的损伤,再灌注开始后即刻应用30 s缺血、30 s再通,三次循环的缺血后处理对骨骼肌的缺血再灌注损伤即有保护作用,不仅减少了骨骼肌再灌注区域中性粒细胞浸润(MPO)和血清氧自由基水平(MDA)水平,而且减少了骨骼肌的死亡程度;再灌注5 min后应用缺血后处理并没有降低骨骼肌缺血再灌注区域的MPO和血清MDA水平,也没有降低骨骼肌缺血再灌注后的死亡程度,与直接缺血再灌注组相同,对骨骼肌缺血再灌注损伤并没有保护作用.结论 骨骼肌缺血后再灌注开始前立刻应用缺血后处理对大鼠骨骼肌缺血再灌注损伤有一定的保护效果,可以减少骨骼肌缺血再灌注损伤后的死亡程度;缺血后处理应用时机非常重要,再灌注5 min后应用缺血后处理则失去对骨骼肌缺血再灌注损伤的保护作用.     

缺血后处理对大鼠移植肝缺血再灌注损伤的保护作用

目的探讨在体条件下缺血后处理对大鼠移植肝缺血再灌注损伤的保护作用及其可能机制。方法采用SD大鼠原位肝移植模型,供肝冷保存时间100min,无肝期控制于18min以内,60只雄性健康SD大鼠随机分为3组,对照组12只,缺血再灌注损伤组和后处理组各24只。对照组开腹后仅游离肝周韧带;缺血再灌注损伤组受体大鼠供肝切除前仅以肝素化生理盐水经门静脉灌注;后处理组供肝植入后完全再灌注前,给予多次短暂复灌复停作为缺血后处理。缺血再灌注损伤组、后处理组受体一半（6只）于再灌注后2h留取血液及肝组织,另一半（6只）于再灌注后6h留取肝组织。对照组于关腹后相应时间留取血液及肝组织。各组分别检测肝功能,采用酶联免疫吸附法测定血清肿瘤坏死因子Or．和中性粒细胞弹性蛋白酶。根据酶促反应原理,利用分光光度仪测定肝脏谷胱甘肽过氧化物酶、丙二醛、髓过氧化物酶、超氧化物歧化酶。肝组织HE染色后光镜下观察组织学变化。结果缺血再灌注损伤组和后处理组血清肝功能指标、炎性细胞因子水平及肝组织过氧化物含量均高于对照组（P〈0．05）,而后处理组较缺血再灌注损伤组则明显低（P〈0．05）;缺血再灌注损伤组和后处理组肝组织抗氧化酶活力显著低于对照组（P〈0．05）,而后处理组较缺血再灌注损伤组则明显高（P〈0．05）。结论缺血后处理对大鼠移植肝的缺血再灌注损伤有明显的保护作用。提高组织的抗氧化能力和降低炎性细胞因子水平可能是缺血后处理保护作用的机制之一。     

Glucagon-like peptide 1 can directly protect the heart against ischemia/reperfusion injury

Glucagon-like peptide 1 (GLP-1), a gut incretin hormone that stimulates insulin secretion, also activates antiapoptotic signaling pathways such as phosphoinositide 3-kinase and mitogen-activated protein kinase in pancreatic and insulinoma cells. Since these kinases have been shown to protect against myocardial injury, we hypothesized that GLP-1 could directly protect the heart against such injury via these prosurvival signaling pathways. Both isolated perfused rat heart and whole animal models of ischemia/reperfusion were used, with infarct size measured as the end point of injury. In both studies, GLP-1 added before ischemia demonstrated a significant reduction in infarction compared with the valine pyrrolidide (an inhibitor of its breakdown) or saline groups. This protection was abolished in the in vitro hearts by the GLP-1 receptor antagonist exendin (9-39), the cAMP inhibitor Rp-cAMP, the PI3kinase inhibitor LY294002, and the p42/44 mitogen-activated protein kinase inhibitor UO126. Western blot analysis demonstrated the phosphorylation of the proapoptotic peptide BAD in the GLP-1-treated groups. We show for the first time that GLP-1 protects against myocardial infarction in the isolated and intact rat heart. This protection appears to involve activating multiple prosurvival kinases. This finding may represent a new therapeutic potential for this class of drug currently undergoing clinical trials in the treatment of type 2 diabetes.     

不同热缺血时间再灌注损伤对大鼠移植胰的影响

目的探讨不同热缺血时间再灌注对大鼠移植胰的损伤情况。方法6只正常SD大鼠为对照组,18只糖尿病SD大鼠随机分为WIR0组(热缺血时间0min,n=6)、WIR15(热缺血时间15min,n=6)和WIR30(热缺血时间30min,n=6)。WIR0组、WIR15组和WIR30组均行胰腺移植。观测各组再灌注前后血糖,再灌注后2h血清TNF-α和NO的含量、移植胰组织中SOD、MPO和MDA含量,组织学变化及细胞凋亡情况。结果(1)WIR0组和WIR15组较再灌注前血糖即下降,再灌注后WIR15组和WIR30组较WIR0组、WIR30组较WIR15组血糖高。(2)再灌注后WIR0组、WIR15组和WIR30组较对照组、WIR15组和WIR30较WIR0组、WIR30较WIR15组血清TNF-α含量高、NO含量低。(3)再灌注后WIR0组与WIR15组和WIR30组较对照组、WIR15组和WIR30组较WIR0组与对照组、WIR30组较WIR15组与对照组胰腺组织中SOD活性低、MDA含量高、MPO活性高。(4)再灌注后2hWIR0组与WIR15组和WIR30组较对照组、WIR15组较WIR0组凋亡指数高。(5)WIR30组供胰组织病理损伤最为严重。结论缺血再灌注损伤可导致移植胰细胞凋亡;在冷缺血180min的条件下,大鼠供胰的最大耐受热缺血时间为15min。     

Use of levosimendan in patients with ischemic heart disease following mechanical reperfusion

Cardiac failure is among the most significant conditions associated with acute coronary syndrome. In ischemic heart disease, serious hemodynamic problems are reported in patients with left ventricular dysfunction during the acute phase despite mechanical revascularization. Several positive inotropic agents in addition to intra-aortic balloon pump (IABP) are required to support patients with impaired left ventricular pump function during this phase. Intravenous inotropic agents, beta-mimetics, and phosphodiesterase inhibitors lead to increases in the incidence of arrhythmia and myocardial O₂ consumption owing to their effect of increasing intracellular calcium amount, although they produce rapid hemodynamic improvements in cardiac failure. This causes severe problems particularly in cardiac failure of ischemic origin. Recently, levosimendan, a calcium-sensitizing agent with cardioprotective properties, is being used alone or in combination with IABP in cases with severe left ventricular systolic dysfunction during mechanical revascularization procedures (percutaneous coronary interventions, coronary bypass surgery). This review includes studies with levosimendan in cases not recovering due to myocardial stunning in the acute phase despite mechanical approaches applied.     

Effects of volatile anesthetics on cardiac metabolism in the low-pressure perfused rat heart

Effects of halothane, enflurane and isoflurane on the myocardial metabolism were studied in the rat heart-lung preparation. Hearts were perfused at a low perfusion pressure (SBP 50mmHg, DBP 30mmHg) with succinate or glutamate as substrates. Thirty minutes after the perfusion, intramyocardial ATP, pyruvate, lactate and glycogen were measured enzymatically. Although there was no significant difference in ATP levels of hearts with either substrate, and whether or not volatile anesthetics were present, 1% halothane and 1.5% isoflurane reduced the L/P ratio when succinate was substrate (24.46 ± 4.81, 17.68 ± 9.10 vs 39.82 ± 10.83), and 2% enflurane decreased it when glutamate was substrate (22.25 ± 10.99 vs 38.44 ± 6.55). The glycogen levels in volatile anesthetics groups were lower than control when succinate was substrate. The improvement of energy demand-supply balance by inhalation anesthetics may be stronger than their inhibition of electron transport in mitochondria under certain ischemic conditions.(Kashimoto S, Hinohara S, Kumazawa T: Effects of volatile anesthetics on cardiac metabolism in the low-pressure perfused rat heart. J Anesth 2: 12–16, 1988)     

缺血后处理对大鼠离体心脏缺血再灌注损伤的作用

目的探讨缺血后处理对大鼠离体心脏缺血再灌注损伤的作用。方法24只Wistar大鼠,随机分为3组(n=8):正常对照组(C组)、缺血再灌注组(I/R组)、缺血后处理组(IPC组)。采用大鼠离体心脏Langendorff灌流模型,C组用K-H液灌注160min;I/R组全心缺血40 min,再灌注120 min; IPC组全心缺血40 min后,再灌注10 s,缺血10 s,反复6次,然后持续再灌注118 min。测定再灌注15、30、120 min时冠脉流量(CF)及冠脉流出液心肌肌钙蛋白I(cTnI)浓度,再灌注120 min时,取心肌组织,测定丙二醛(MDA)含量、超氧化物歧化酶(SOD)活性,电镜下观察心肌细胞超微结构。结果缺血再灌注可导致CF降低,冠脉流出液cTnI浓度升高,心肌SOD活性下降,MDA含量升高,心肌细胞超微结构产生病理学改变,缺血后处理可减弱上述改变。结论缺血后处理减轻脂质过氧化反应,对大鼠离体缺血再灌注心脏产生保护作用。     

Interleukin-6 and STAT3 protect the liver from hepatic ischemia and reperfusion injury during ischemic preconditioning

BACKGROUND: Ischemic preconditioning has been shown to protect the liver from ischemia/reperfusion injury. We hypothesized that IL-6 directly modulates the protective effects of ischemic preconditioning. METHODS: Three weeks after undergoing splenic transposition, wild-type C57BL/6 and IL-6 null mice underwent 75 minutes of total hepatic ischemia with or without prior ischemic preconditioning (10 minutes of ischemia followed by 15 minutes of reperfusion). After reperfusion, serum ALT, serum IL-6, hepatic IL-6 mRNA, hepatic pSTAT3, and liver histology were evaluated. RESULTS: In wild-type mice, survival at 24 hours was greater in the preconditioned group compared with the non-preconditioned group (75% vs 40%, P<.05). In IL-6 null mice, however, ischemic preconditioning did not improve survival when compared with the non-preconditioned group. Preconditioning significantly reduced hepatocellular injury in wild-type mice (P<.05) when compared with IL-6 null animals. This protection was associated with significant increases in serum IL-6, hepatic IL-6 mRNA, and hepatic pSTAT3 levels (P<.05). The protective effects of ischemic preconditioning that correlated with significant increases in systemic IL-6, hepatic IL-6 mRNA abundance, and pSTAT3 levels, were not observed in IL-6 null mice. CONCLUSIONS: The protective effects of ischemic preconditioning during total hepatic ischemia/reperfusion injury are dependent on IL-6 signaling and are associated with increased phosphorylation of hepatic STAT3.