Ischemic Postconditioning Group Research Articles

Ischemic postconditioning protects cardiomyocytes against ischemia/reperfusion injury by inducing MIP2

Cardiomyocytes can resist ischemia/reperfusion (I/R) injury through ischemic postconditioning (IPoC) which is repetitive ischemia induced during the onset of reperfusion. Myocardial ischemic preconditioning up-regulated protein 2 (MIP2) is a member of the WD-40 family proteins, we previously showed that MIP2 was up-regulated during ischemic preconditioning (IPC). As IPC and IPoC engaged similar molecular mechanisms in cardioprotection, this study aimed to elucidate whether MIP2 was up-regulated during IPoC and contributed to IPoC-mediated protection against I/R injury. The experiment was conducted on two models, an in vivo open chest rat coronary artery occlusion model and an in vitro model with H9c2 myogenic cells. In both models, 3 groups were constituted and randomly designated as the sham, I/R and IPoC/hypoxia postconditioning (HPoC) groups. In the IPoC group, after 45 min of ischemia, hearts were allowed three cycles of reperfusion/ischemia phases (each of 30 s duration) followed by reperfusion. In the HPoC group, after 6 h of hypoxia, H9c2 cells were subjected to three cycles of 10 minute reoxygenation and 10 minute hypoxia followed by reoxygenation. IPoC significantly reduced the infarct size, plasma level of Lactate dehydrogenase and creatine kinase MB in rats. 12 h after the reperfusion, MIP2 mRNA levels in the IPoC group were 10 folds that of the sham group and 1.4 folds that of the I/R group. Increased expression of MIP2 mRNA and attenuation of apoptosis were similarly observed in the HPoC group in the in vitro model. These effects were blunted by transfection with MIP2 siRNA in the H9c2 cells. This study demonstrated that IPoC induced protection was associated with increased expression of MIP2. Both MIP2 overexpression and MIP2 suppression can influence the IPoC induced protection.

Comparison of cardioprotective and anti-inflammatory effects of ischemia pre- and postconditioning in rats with myocardial ischemia–reperfusion injury

To compare cardioprotective and anti-inflammatory effects of ischemia preconditioning (IPC) and ischemia postconditioning (IPOC) in a rat myocardial ischemia-reperfusion injury (IRI) model. Forty Sprague-Dawley rats were randomly divided equally into four groups. In the groups other than the sham group, the left anterior descending coronary artery was ligated for 30 min followed by a 180 min reperfusion in vivo. The control group was subjected to no additional intervention, the IPC group to three cycles of 5 min ischemia separated by 5 min reperfusion before the index ischemia and the IPOC group to three cycles of 10 s ischemia separated by 10 s reperfusion immediately after the end of the index ischemia. Hemodynamic changes during the ischemia and reperfusion were recorded. At 180 min of reperfusion, serum concentrations of troponin I (TnI), tumor necrosis factor α (TNF-α) and high-mobility group box 1 (HMGB-1) were assayed, and the infarction size was assessed by Evans blue and triphenyltetrazolium chloride staining. Compared to the control group, infarct size and serum concentrations of TnI, TNF-α and HMGB1 at 180 min of reperfusion were significantly reduced in the IPC and IPOC groups. However, infarct size and serum concentrations of TNF-α and HMGB1 at 180 min of reperfusion were significantly increased in the IPOC group compared to the IPC group. In the rats with myocardial IRI in vivo, both IPC and IPOC can produce significant cardioprotective and anti-inflammatory effects. However, cardioprotective and anti-inflammatory effects provided by IPOC are weaker than with IPC.

Ischemic Postconditioning Attenuates Lung Reperfusion Injury and Reduces Systemic Proinflammatory Cytokine Release Via Heme Oxygenase 1

Systemic inflammatory response following ischemia-reperfusion injury (IRI) to a specific organ may cause injuries in multiple remote organs. The emergence of ischemic postconditioning (IPO) provides a potential method for experimentally and clinically attenuating various types of organ postischemic injuries. We have shown that IPO can attenuate lung IRI by up-regulating the protein expression of heme oxygenase-1(HO-1). This study tested the hypothesis that IPO attenuates systemic inflammatory responses following lung IRI by activating HO-1. Anaesthetized and mechanically ventilated adult Sprague-Dawley rats were randomly assigned to one of the following groups (n = 8 each): the sham-operated control group, the ischemia-reperfusion (IR) group (40 min of left-lung ischemia and 120 min of reperfusion), the IPO group (three successive cycles of 30-s reperfusion per 30-s occlusion before restoring full perfusion), and the zinc protoporphyrin IX (ZnP) plus IPO group (ZnP, an inhibitor of HO-1, was injected intraperitoneally at 20 mg/kg 24 h prior to the experiment, and the rest of the procedures were similar to that of the IPO group). Lung injury was assessed by arterial blood gas analysis, wet-to-dry lung weight ratio and tissue histologic and biochemical changes. The lung tissue and plasma levels of lipid peroxidation were determined by measuring the contents of malondialdehyde (MDA) production. Protein expression of HO-1 was determined by Western blotting. Pulmonary neutrophil was counted. Lung tissue myeloperoxidase (MPO) activity as well as plasma levels of proinflammatory cytokines tumor necrosis factor-α (TNF-α), interleukines 6 and 8 (IL-6, IL-8) were determined by spectrophotography. Lung ischemia-reperfusion led to severe lung pathologic morphologic changes and increased pulmonary MDA production, neutrophil count, and MPO activity and reduced arterial oxygen partial pressure (all P < 0.05 IR versus sham), accompanied with a compensatory increase in HO-1 protein and activity. Plasma levels of TNF-α, IL-6, and IL-8 were increased in the IR group (all P < 0.05 versus sham). IPO attenuated or prevented all the above changes, except that it further increased lung HO-1 activity. Treatment with ZnP abolished all the protective effects of postconditioning. Postconditioning attenuated pulmonary neutrophil accumulation and activation and lung IRI and reduced systemic inflammatory responses by activating HO-1.

Synergistic protection of Danhong Injection (丹红注射液) and ischemic postconditioning on myocardial reperfusion injury in minipigs

To explore the synergistic protection of Danhong Injection (丹红注射液, DHI) and ischemic postconditioning on myocardial reperfusion injury in minipigs. Acute myocardial infarction model was made by balloon occlusion in left anterior descending coronary artery (LAD) of minipigs, and then postconditioning was simulated through inflation/deflation of the angioplasty balloon. Minipigs were divided into four groups: the sham operation group (SH group), the ischemia/reperfusion group (I/R group), the ischemic postconditioning group (POC group) and DHI combined with ischemic postconditioning group (PAD group, DHI 20 mL through ear vein), six in each group. After 24-h continuous observation, myocardial infarction size was assessed by triphenyltetrazolium staining (TTC). Morphological changes of ischemic myocardium were observed by light microscopy, and cardiomyocyte ultrastructure was studied with electron microscopy. The superoxide dismutase (SOD) and malondialdehyde (MDA) activity in heart homogenates were measured by a biochemical method. The myocardial infarction size was smaller in the POC group than in the I/R group (0.26 ± 0.02 vs. 0.37 ± 0.09, P<0.05), and the PAD group (0.14 ± 0.08) displayed a significantly reduced infarction size relative to the I/R group (P<0.01) and POC group (P<0.05). The damage of myocardial tissue was severe in the I/R group shown by light and electron microscopy: myocardial fibers disorder, sarcoplasmic dissolution, myofilament fracture, mitochondria swelling and even vacuolization formation and a large number of inflammatory cell infiltrations. Compared with the I/R group, reduction of reperfusion injury in the PAD group included more orderly arranged myocardial fibers, less infiltration of inflammatory cells and maintenance of mitochondrial integrity. Compared with the I/R group, the damage of myocardial tissue in the POC group was improved, but not as significant as that in the PAD group. SOD levels in the POC group and the PAD group were significantly higher than those in the I/R group (96.96 ± 13.43, 112.25 ± 22.75 vs. 76.32 ± 10.63, P<0.05), and MDA was significantly lower in the POC group and the PAD group compared to the I/R group (1.27 ± 0.19, 1.09 ± 0.21 vs. 1.47 ± 0.16, P<0.05). DHI and ischemic postconditioning show a synergistic cardioprotection on myocardial reperfusion injury in minipigs.

Effects of penehyclidine hydrochloride postconditioning on hind limb ischemia-reperfution injury in rats

Objective To investigate the effects of penehyclldine hydrochloride postconditioning on hind limb ischemia-reperfusion （I/R） injury in rats. Methods One hundred and forty-four male Wistar rats weighing 220-250 g were randomly divided into 4 groups （n = 36 each）： control group （group C）, I/R group, ischemic postconditioning group （group IPO） and penehyclidine hydrochloride postconditioning group （group PHPO）. Limb ischemia was induced by occlusion of bilateral hind limb for 3 h followed by 24 h reperfusion. In group IPO, the animals were subjected to three cycles of 30 s reperfusion and 30 s ischemia on the proximal part of the hind limb,while in group PHPO, the animals received iv injection of penehyclidine hydrochloride 0.15 mg/kg （in normal saline 1 wl） via the caudal vein immediately after reperfusion. Blood samples from the inferior vena cava were taken for determination of activities of serum LDH and CK and concentrations of TNF-α and IL-10 at O, 1, 3, 6, 12 and 24 h of reperfusion. Then 6 rats were sacrificed at each time point respectively and the quadriceps femoris tissues of the hind limb were removed for determination of MDA content, activities of SOD, MPO, Na＋ -K＋-ATPase and Ca2 ＋ - Mg2＋ -ATPase, and HIF- 1 α expression and microscropic examination. Results Compared with group C, the activities of serum LDH and CK and concentrations of TNF-α and IL-10 were significantly increased, the activities of SOD, Na＋-K＋-ATPase and Ca2＋-Mg2＋-ATPase were decreased, MPO activities and MDA content were increased, HIF-1α expression was up-regulated in group I/R, IPO and PHPO （P 〈0.05 or 0.01）. Compared with group I/R, the serum activities of serum LDH and CK and TNF-α concentration were significantly decreased, IL- 10concentration was increased, activities of SOD, Na＋ -K＋ -ATPase and Ca2＋ -Mg2＋ -ATPase were increased, and MPO activity and MDA content were decreased, HIF-1α expression was down-regulated （P 〈 0.01）, and the pathologic changes was reduced in group IPO and PHPO. Compared with group IPO, the serum LDH activity and TNF-α concentration were significantly decreased, CK activity and IL-10 concentration were increased, activities of SOD and MPO and MDA content were decreased and HIF-1α expression was down-regulated （P 〈 0.05 or 0.01）.Conclusion Penehyclidine hydrochloride postconditioning can reduce hind limb I/R injury in rats, and the mechanism is related to inhibiting inflammatory response and lipid peroxidation, reducing calcium overload and improving cellular energy metabolism. Key words: Quinuclidines; Reperfusion injury; Extremities

Effects of ischemic preconditioning versus ischemic postconditioning on myocardial ischemia-reperfusion-induced inflammatory response in rats

Objective To compare the effects of ischemic preconditioning versus ischemic postconditioning on myocardial ischemia-reperfusion （I/R）-induced inflammatory response in rats. Methods Forty male SD rats weighing 290-320 g were randomly divided into 4 groups （ n = 10 each）： Ⅰ group sham operatin （group S）; Ⅱ group I/R; Ⅲ group ischemic preconditioning （group IPC） and Ⅳ group ischemic postconditioning （group IPOC）.Myocardial I/R was induced by 30 min ligation of left anterior descending branch （LAD） of coronary artery followed by reperfusion. In group IPC myocardial I/R was preceded by 3 cycles of ischemia followed by reperfusion （each lasting 5 min） while in group IPOC 3 cycles of I/R （each lasting 10 s） was started at the end of 30 min myocardial ischemia. MAP, HR and RPP （ MAP × HR） were recorded before （baseline） and at 1 and 20 min of ischemia and 60, 120 and 180 min of reperfusion. Venous blood samples were collected at 30 and 180 min of reperfusion for determination of serum concentrations of TNF-α, IL-6, high-mobility group box 1 （HMGB1） and cTnI. The animals were sacrificed at 180 min of reperfusion and the myocardial infarct size was measured. Results Myocardial I/R significantly decreased MAP and RPP and increased myocarcial infarct size, serum concentrations of TNF-α, IL-6,HMGB1 and cTnI in group I/R as compmed with group S. Ischemic pre- and postconditioning significantly increased MAP and reduced myocardial infarct size and I/R-induced increase in serum TNF-α, HMGB1 and cTnI concentrations in group Ⅲ and Ⅳ as compared with group Ⅱ （I/R）. The myocardial infarct size was significantly larger and the serum concentrations of TNF-α, IL-6 and HMGB1 were significantly higher in ischemic postconditioning group than in the preconditioning group. Conclusion Ischemic preconditioning is more effective in attenuating the myocardial I/R-induced inflammatory response than the ischemic postconditioning. Key words: Ischemic preconditioning, myocardial; Myocardial repcrfusion injury; Inflammation; Ischemia postconditioning

E0042 The effect of ischaemic postconditioning on the structure, function and Cx43 of mitochondria in rabbit myocardial isehemia/reperfusion injury

ObjectiveTo investigate the effects of ischaemic postconditioning on structural and functional and connexin 43(Cx43) changes of mitochondria induced by myocardial ischaemia reperfusion (IR) injury of rabbits in vivo and potential...

Ischemic Postconditioning Inhibits Apoptosis aft er Acute Myocardial Infarction in Pigs

Recent studies have shown that ischemic postconditioning reduces myocardial ischemia-reperfusion (I/R) injury; however, the effects of inhibiting apoptosis on cardioprotection induced by ischemic postconditioning remain to be determined. The objective of this study was to investigate whether ischemic postconditioning attenuates myocardial I/R injury by reduced apoptosis in a closed-chest pig model of acute myocardial infarction. Diannan small-ear pigs were randomly divided into 3 groups (5/group): (1) The sham group underwent a sham operation without ischemia; (2) the I/R group received 60 minutes of ischemia and 72 hours of reperfusion; and (3) the ischemic postconditioning (Postcond) group was treated the same as the I/R group except that the pigs received 8 cycles of 30 seconds of reperfusion and 30 seconds of ischemia at the onset of reperfusion. After 72 hours of reperfusion, infarct size was measured by 2,3,5-triphenyltetrazolium chloride staining. Apoptotic cells in the peri-infarct myocardium were evaluated with the terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) method, and apoptosis-related molecules were studied with western blotting analysis. After 72 hours of reperfusion, mean (±SEM) infarct size was significantly smaller in the Postcond group than in the I/R group (23.26% ± 3.13% versus 10.89% ± 2.02%, P < .05). Apoptotic myocytes in the peri-infarct region were lower in the Postcond group than in the I/R group (15.31% ± 4.58% versus 33.83% ± 4.44%, P < .05). This decrease in the extent of apoptosis was accompanied by a significant decrease in Bax expression (0.306 ± 0.075 versus 0.433 ± 0.102 for the I/R group; P < .05) and a significant increase in Bcl-2 expression (1.801 ± 0.227 versus 1.267 ± 0.308 for the I/R group; P < .05). In a clinically relevant closed-chest pig model of myocardial infarction, these data suggest the following: (1) Ischemic postconditioning reduces infarct size following prolonged reperfusion, and (2) this cardioprotective effect is likely achieved via antiapoptotic mechanisms.

福辛普利钠预处理对缺血后适应大鼠心肌组织Toll样受体表达及炎症因子的影响

To observe the effects of ischemic post-conditioning (IPoC) on Toll-like receptor expression and myocardial inflammatory infiltration in myocardial tissue of rats with ischemic reperfusion (I/R) injury and fosinopril sodium pre-treatment, a total of 60 Sprague-Dawley rats were randomly assigned to the following groups: sham (suture around left anterior descending coronary artery was not tied), ischemic reperfusion (30 min<italic> in situ</italic> transient occlusion of the left anterior descending artery, followed by 1 h reperfusion), IPoC (30-min occlusion of left anterior descending artery, followed by three cycles of 10-s reperfusion/10-s ischemia prior to 1-h reperfusion), and fosinopril sodium +IPoC (fosinopril sodium pre-treatment, 0.9 mg/kg, for 14 d, followed by 2-h I/R after last gavage, and IPoC treatment during I/R). Arterial blood and heart samples were extracted after 1-h reperfusion. Serum cTnT levels were measured using the colorimetric method, myocardial infarct size was measured using nitrotetrazolium blue/chloride staining, TNF-α and MCP-1 levels in myocardial tissue were measured by ELISA. TLR2 and TLR4 expression in myocardial tissue was analyzed using immunohistochemistry, and infiltrating cells were detected by hematoxylin-eosin staining. Compared with the I/R group, myocardial enzyme cTnT levels and infarct size significantly decreased in the IPoC group (<italic>P</italic> < 0.01). In addition, MCP-1 and TNF-α levels, as well as the number of infiltrating cells in myocardial tissue, significantly decreased (<italic>P</italic> < 0.05, <italic>P</italic> < 0.01, respectively). TLR2 and 4 expressions also significantly decreased in the IPoC group (<italic>P</italic> < 0.05, <italic>P</italic> < 0.01, respectively). Compared with the IPoC group, the combination of fosinopril sodium and IPoC further reduced infarct size (<italic>P</italic> < 0.05), as well as TNF-α levels in myocardial tissue (<italic>P</italic> < 0.01). TLR2 and 4 expressions and infiltration of inflammatory cells were also significantly decreased (<italic>P</italic> < 0.05). Therefore, fosinopril sodium enhanced the protective effect of IPoC in a rat model of myocardial I/R injury. These mechanisms could be related to inhibition of the TLR signal pathway and chemotactic factor reactions.

Examination of Protective Effect of Ischemic Postconditioning After Small Bowel Autotransplantation

Ischemia/reperfusion (I/R) injury is a serious condition that results from some surgical procedures, including intestinal transplantation. Ischemic postconditioning is defined as brief periods of reperfusion alternating with reocclusion applied during the early minutes after reperfusion. The objective of this study was to investigate the effect of ischemic postconditioning before small bowel autotransplantation. Total orthotopic intestinal autotransplantation was performed in 30 white domestic pigs. Grafts were stored in cold University of Wisconsin solution for 1, 3, or 6 hours. Duration of reperfusion was 3 hours in all grafts. Before reperfusion, the intestine was postconditioned via 3 cycles of ischemia for 30 seconds and reperfusion for 30 seconds (ischemic postconditioning protocol). Tissue from the small intestine was obtained after laparotomy (control group) and at the end of reperfusion periods. To monitor oxidative stress, tissue concentrations of malondialdehyde and reduced glutathione, and activity of superoxide dismutase were determined at spectrophotometry. Tissue damage on sections stained with hematoxylin- eosin was evaluated using a quantitative method (Scion Image software; Scion Corp, Frederick, Maryland). Our results demonstrated that ischemic postconditioning significantly decreased the reperfusion-ended lipid peroxidation value (mean ± SEM, 142.0 ± 7.1 μmol/g vs 125.0 ± 2.1 μmol/g; P < .05). Moreover, the capacity and activity of endogenous antioxidant protective systems (glutathione 789 ± 8.0 μmol/g vs 934 ± 5.7 μmol/g, and superoxide dismutase 110 ± 9 IU/g vs 126 ± 4 IU/g; P < .05) remained higher in the ischemic postconditioning groups compared with tissues without ischemic postconditioning. At quantitative analysis, tissue injury was increased by the duration of cold preservation. The greatest injury was observed in the mucosal and submucosal layers and in the depth of crypts after 6 hours of preservation. Ischemic postconditioning significantly decreased intestinal wall injury in each group ( P < .05). It was concluded that ischemic postconditioning before reperfusion mitigated oxidative stress and histologic damage during small bowel autotransplantation.

Roles of reactive oxygen species and phosphatidyl-inositol 3- kinase-Akt in neuroprotection against spinal cord ischemia-reperfusion injury by ischemic postconditioning or controlled low perfusion pressure in rats

Objective To investigate the roles of reactive oxygen species (ROS) and phosphatidyl-inositol 3-kinase-Akt (PI3K-Akt) in neuroprotection against spinal cord ischemia-reperfusion (I/R) injury by ischemic postconditioning (IP) or controlled low perfusion pressure (LR). Methods One hundred and twenty-six adult male SD rats weighing 300-350 g were randomly divided into 7 groups (n = 18 each): group Ⅰ I/R; group ⅡIP; group Ⅲ LR; group Ⅳ IP + LY-294002 (IP + LY); group Ⅴ LR + LY-294002 (LR + LY); group Ⅵ IP +N-acetylcysteine (IP+ N) and group Ⅶ LR+ N-acetyl-cysteine (LR+ N). Spinal cord ischemia was induced by 9 min occlusion of the thoracic aorta combined with controlled hypotension (MAP = 40 mm Hg). In IP group the animals were subjected to 3 cycles of 30 s reperfusion interspersed with 30 s ischemia immediately after release of aortic occlusion. In LR group MAP was maintained at 40 mm Hg for another 5 min immediately after release of aortic occlusion, then increased to 100 mm Hg. In group Ⅳ-Ⅶ LY-294002 (PI3K inhibitor) 25 mg/kg or N-acetylcysteine (ROS scavenger) 100 mg/kg were administered at the onset of reperfusion. Twelve animals in each group were killed at 2 h of reperfusion. The lumbar segment of the spinal cord was removed for determination of the level of Akt phosphorylation and opening of mitochondrial permeability transition pore (mPTP). Neurological function was assessed and scored (15 = normal function, 0 = unable to move hind limb) at 4, 12, 24 and 48 h of reperfusion in another 6 animals in each group. The animals were then killed after last assessment and the lumbar segment of the spinal cord was removed for detection of apoptotic neurons. Results Compared with I/R group,both IP and LR significantly enhanced the level of Akt phosphorylation in the spinal cord, inhibited mPTP opening and neuronal apoptosis and increased neurological function scores. There was no significant difference in the protective effects exerted by IP and LR. The neuroprotective effects exerted by IP and LR were abolished by LY-294002 and N-acetylcysteine. Conclusion Ischemic postconditioning or controlled low perfusion pressure can protect the spinal cord from I/R injury by activating PI3K-Akt and inhibiting mitochondrial permeability. Key words: Reactive oxygen species; 1-Phosphatidylinositol 3-kinase; Protein kinases; Perfusion; Reperfusion injury; Spinal cord; Ischemic postconditioning

Mechanisms of Chinese herbs combined with ischemic postconditioning in protecting myocardium of rats from ischemia-reperfusion injury

To observe the effects of Chinese patent medicines with the function of replenishing qi to activate blood (RQAB) plus ischemic postconditioning (IPoC) in protecting myocardium of rats from ischemia-reperfusion (I/R) injury, and to explore the possible mechanisms. Seventy-five Sprague-Dawley rats were randomly divided into sham-operated group (the suture was penetrated around the left anterior descending coronary artery, but not tied, n=15), I/R group (30 minutes of in situ transient occlusion of the left anterior descending artery, followed by 1 hour of reperfusion, n=15), IPoC group (30 minutes occlusion of the left anterior descending artery, followed by 3 cycles of 10 s of reperfusion/10 s of ischemia before 1-hour reperfusion, n=15), RQAB plus IPoC group (pretreated with 0.162 g/kg Xinyue Capsule and 0.135 g/kg Xiongshao Capsule for 14 days, and treated with IPoC 2 h after the final gavage, n=15), fosinopril sodium plus IPoC group (pretreated with fosinopril sodium, 0.9 mg/kg, n=15). Serum creatine kinase-MB (CK-MB) activity and cardiac troponin T (cTnT) level were detected; myocardial infarction size was measured by nitrotetrazolium blue chloride staining; Toll-like receptor 2 (TLR2) and TLR4 in myocardial tissue were examined by immunohistochemical method; interleukin-1beta (IL-1beta) and interleukin-6 (IL-6) levels in myocardial tissue were examined by enzyme-linked immunosorbant assay. Compared with the I/R group, myocardial enzymes and infarction size were decreased significantly in the IPoC group (P<0.01); expressions of TLR2 and TLR4 and levels of IL-1beta and IL-6 in myocardial tissues were also significantly lower than those in the I/R group (P<0.05). Compared with the fosinopril sodium plus IPoC group, expressions of TLR2 and TLR4 were decreased significantly in the RQAB plus IPoC group (P<0.05, P<0.01). Compared with IPoC, RQAB plus IPoC reduced the infarction size and the release of myocardial enzyme CK-MB (P<0.01), and decreased the expressions of TLR2 and TLR4 and the levels of IL-1beta and IL-6 (P<0.05, P<0.01) in myocardial tissues. Pretreatment with Chinese herbs for nourishing qi and activating blood circulation can enhance the protective effect of IPoC on rat myocardial I/R injury, and its mechanism may be related to inhibition of TLR expression and expressions of the downstream proinflammatory cytokines.

Role of matrix metalloproteinase‐2 in the cardioprotective effect of ischaemic postconditioning

The activation of matrix metalloproteinases (MMPs) contributes to myocardial injury at the onset of reperfusion; however, their role in ischaemic postconditioning is unknown. The aim of the present study was to examine the effects of ischaemic postconditioning on MMP activity in isolated rabbit hearts. The isolated rabbit hearts were subjected to 30 min of global ischaemia followed by 180 min of reperfusion (I/R group; n = 8). In the ischaemic postconditioning group (n = 8), a postconditioning protocol was performed (2 cycles of 30 s reperfusion-ischaemia). In other experiments, we added doxycycline, an MMP inhibitor, at 25 (n = 7) or 50 micromol l(1) (n = 8) during the first 2 min of reperfusion. Coronary effluent and left ventricular tissue were collected during pre-ischaemic conditions and at different times during the reperfusion period to measure MMP-2 activity and cardiac protein nitration. We evaluated ventricular function and infarct size. In the I/R group, infarct size was 32.1 +/- 5.2%; Postcon reduced infarct size to 9.5 +/- 3.8% (P < 0.05) and inhibited MMP-2 activity during reperfusion. The administration of doxycycline at 50 micromol l(1) inhibited MMP-2 activity and cardiac protein nitration and reduced the infarct size to 9.7 +/- 2.8% (P < 0.05). A lower dose of doxycycline (25 micromol l(1)) failed to inhibit MMP-2 activity and did not modify the infarct size. Our results strongly suggest that ischaemic postconditioning may exert part of its cardioprotective effects through the inhibition of MMP-2 activity.

Ischemic postconditioning may not influence early brain injury induced by focal cerebral ischemia/reperfusion in rats

BackgroundExperimental studies have shown that ischemic postconditioning can reduce neuronal injury in the setting of cerebral ischemia, but the mechanisms are not yet clearly elucidated. This study was conducted to determine whether ischemic postconditioning can alter expression of heat shock protein 70 and reduce acute phase neuronal injury in rats subjected to transient focal cerebral ischemia/reperfusion.MethodsFocal cerebral ischemia was induced by intraluminal middle cerebral artery occlusion for 60 min in twenty male Sprague-Dawley rats (250-300 g). Rats were randomized into control group and an ischemic postconditioning group (10 rats per group). The animals of control group had no intervention either before or after MCA occlusion. Ischemic postconditioning was elicited by 3 cycles of 30 s reperfusion interspersed by 10 s ischemia immediately after onset of reperfusion. The infarct ratios, brain edema ratios and motor behavior deficits were analyzed 24 hrs after ischemic insult. Caspase-3 reactive cells and cells showing heat shock protein 70 activity were counted in the caudoputamen and frontoparietal cortex.ResultsIschemic postconditiong did not reduce infarct size and brain edema ratios compared to control group. Neurologic scores were not significantly different between groups. The number of caspase-3 reactive cells in the ischemic postconditioning group was not significantly different than the value of the control group in the caudoputamen and frontoparietal cortex. The number of cells showing heat shock protein 70 activity was not significantly different than the control group, as well.ConclusionsThese results suggest that ischemic postconditioning may not influence the early brain damage induced by focal cerebral ischemia in rats.

Effects of combination of ischemic postconditioning and remote ischemic postconditioning on focal cerebral ischemia-reperfusion injury in rats

Objective To investigate the effects of combination of ischemic postconditioning (IPC) and remote IPC on the focal cerebral ischemia-reperfusion (I/R) injury in rats. Methods Eighty-six adult male SD rats were randomly divided into 5 groups: sham operation group (group Ⅰ, n = 19) , I/R group (group Ⅱ, n=19) , IPC group (group Ⅳ, n = 16) , remote>IPC group (group Ⅳ, n = 16), and IPC + remote IPC group (group V , n = 16) . Focal cerebral I/R was produced by occlusion of the right middle cerebral artery (MCA) for 1.5 h followed by 24 h of reperfusion. In group Ⅰ, operation was performed but focal cerebral I/R was not produced. In group Ⅲ , the rats were subjected to 3 episodes of 30 s ischemia at 30 s intervals before 24 h of reperfusion. In group Ⅳ, after 1.5 h of the right MCA occlusion, remote IPC was performed by occluding the right femoral artery for 5 min before 24 h of reperfusion. In group Ⅴ, IPC and remote IPC were performed as described in group Ⅲ and Ⅳ respectively before 24 h of reperfusion. The neurologic deficit scores (NDSs) were measured at 2 and 24 h of reperfusion. Ten rats in each group were decapitated at 24 h of reperfusion and brains rapidly . removed for measuring the cerebral infarct size (IS). The expression of microtubule-associated protein 2 (MAP2) were detected. The animals received iv injection of fluorescein isothiocyanate (FICT)-dextran via the femoral vein 1 min before decapitation, for laser confocal scanning. The contralateral side was served as control. Results As compared with group Ⅰ, the NDSs and cerebral IS on the ischemic side were significantly increased, and the cerebral expression of MAP2, plasma volume of the cerebral tissues, and diameter and length of the cerebral microvessel on the ischemic side significantly decreased in group Ⅱ, Ⅲ , Ⅳ and Ⅴ ( P < 0.05 ) . As compared with group Ⅱ, the NDSs and cerebral IS on the ischemic side were significantly decreased, and the cerebral expression of MAP2, plasma volume of the cerebral tissues, and diameter and length of the cerebral microvessel on the ischemic side significantly increased in group ID , Ⅳ and Ⅴ( P < 0.05). Conclusion Although the combination of IPC and remote IPC can attenuate the focal cerebral I/R injury in rats, the efficacy is similar to that of either of them alone. Key words: Reperfusion injury; Brain; Ischemia postconditioning

Proteomics of Ischemia/Reperfusion Injury in Rat Intestine With and Without Ischemic Postconditioning

Intestinal ischemia/reperfusion (I/R) injury is a critical condition associated with high morbidity and mortality. Our previous study showed that ischemic postconditioning (IPo) protects the intestinal mucosa from I/R injury. However, the precise molecular mechanisms of this event remain poorly elucidated. The aim of this study was to investigate the differentially expressed proteins of intestinal mucosa after intestinal I/R with or without IPo, and to explore the potential mechanisms of intestinal I/R injury and the protective effect of IPo in relation to the differential proteins. Intestinal I/R injury was established by occluding the superior mesenteric artery (SMA) for 60 min followed by 60 min reperfusion. The rats were randomly allocated into one of three groups based upon the intervention (n = 8); sham : sham surgical preparation including isolation of the SMA without occlusion was performed; injury: there was no intervention either before or after SMA occlusion; IPo: three cycles of 30 s reperfusion-30 s reocclusion were imposed immediately upon reperfusion. A comparative proteomics approach with two-dimensional gel electrophoresis was used to isolate proteins in intestinal mucosa, the expression of which were regulated by I/R injury post-treated with or without IPo. The differentially displayed proteins were identified through matrix-assisted laser desorption/ionization-time of flight-mass spectrometry (MALDI-TOF-MS). Image analysis revealed that an average of 1300 protein spots were detected on each gel; 16 and 9 proteins showing more than 1.5-fold difference were identified between the Sham versus Injury group and injury group versus IPo group, respectively. The identified proteins were functionally involved in the cellular processes of energy metabolism, anti-oxidation, and anti-apoptosis. This study provided new clues for understanding the mechanisms of IPo against intestinal I/R injury.

Effects of ischemic postconditioning on the expression of heme oxygenase-1 in the liver graft with ischemia and reperfusion injury in rats

To investigate the effects of ischemic postconditioning on the expression of HO-1 in the liver graft ischemia and reperfusion injury in rats. Fifty-six male SD rats were randomly divided into four groups: sham-operation group (sham) (n = 8), ischemia and reperfusion group (I/R)(n = 16), ischemic postconditioning group (IPo) (n = 16) and inhibitor of HO-1 group (ZnPP) (n = 16). Donor livers were preserved in 0 - 4 degrees C normal saline, and the period of cold preservation and anhepatic phase were 90 min and 15 min. At 6 h after portal vein reperfusion, blood samples were obtained from the abdominal aorta to determine the level of serum alanine aminotransferase (ALT), aspartate aminotransferase (AST), simultaneously liver tissues were taken to determine the level of malondialdehyde (MDA), superoxide dismutase (SOD) and heme oxygenase-1 (HO-1) mRNA. The changes of liver tissues were observed by HE staining and electron microscope. SOD activity was significantly lower whereas MDA content was significantly higher in I/R group than that in Sham group (P < 0.05). The expression of HO-1 in I/R group was higher than that in Sham group (P < 0.05). MDA content was significantly lower whereas SOD activity was significantly higher in IPO group than that in I/R group (P < 0.05), and the expression of HO-1 in IPO group was significantly stronger than that in I/R group (P < 0.05). SOD activity and the expression of HO-1 were significantly lower whereas MDA content was significantly higher in ZnPP group than that in I/R group (P < 0.05). The changes of liver tissues also proved the previous results. Ischemic postconditioning attenuates liver graft injury induced by I/R in rats. The mechanism might be related with the induction of HO-1 and enhancement of liver graft antioxidation.

Proteomics study in ischemic postconditioning after prolonged ischemia for lessening the ischemia/reperfusion injury of intestinal in rats

Objective To investigate the changes of proteins expressions in intestinal mucosa of rats after is chemic postconditioning (IPo) against intestinal ischemic/reperfusion (Ⅱ/R) injury of intestine in order to elucidate its potential mechanisms of protective role. Methods Sixteen SD rats were randomly divided into Ⅱ/R group and IPo group ( n = 8). Rats of both groups received an episode of ischemic/reperfusion insult to intstine that was made by occlusion of the superior mesenteric artery (SMA) for 60 minutes. Rats of IPo group underwent three additional episodes of clamping SMA on for 30 seconds and off for 30 seconds successively after prolonged reperfusion/reperfusion of intestine. The intestinal mucosa was taken by scratching immediately after reperfusion in both groups, and total proteins were separated by immobilized pH gradient (IPG) based two-dimensional gel electrophoresis (2-DE). The differentially expressed proteins were analyzed using Image Master 2D Elite 5.0 image analysis software, and the proteins were cut out from the gel and then identified using MALDI-TOF-MS. The biological information of these proteins was looked for in the database of these peptide mass finger-printing (PMF) .Results Ten differentially expressed proteins were found, of which 6 were up-regulated and 4 were down-regulated in IPo group. Nine proteins were identified and characterized by their bioelements including aldose reductase and aldehyde dehydrogenase that were related to anti-oxidative stress and inhibition of cell apoptosis. Conclusions The well-reproducible 2-DE profiles of intestinal mucosa in II/R and IPo groups were established. The potentially protective effects of IPo may be attributed to up-regulating protein expressions of aldose reductase and aldehyde dehydrogenase, and thereby suppressing oxidative stress and cell apoptosis. Key words: Ischemic postconditioning; Intestinal ischemic/reperfusion injury; Proteomics; Aldose reductase; Aldehyde dehydrogenase

Ischaemic post-conditioning protects lung from ischaemia–reperfusion injury by up-regulation of haeme oxygenase-1

Objective The emergence of ischaemic post-conditioning (IPO) provides a potential method for experimentally and clinically attenuating various types of organ injuries. There has been little work, however, examining its effects in the setting of lung ischaemia reperfusion (IR). The stress protein, haeme oxygenase-1 (HO-1), has been found to exert a potent, protective role in a variety of lung injury models. In this study, we hypothesised that the induction of HO-1 by IPO plays a protective role against the deleterious effects of IR in the lung. Methods Anaesthetised and mechanically ventilated adult Sprague–Dawley rats were randomly assigned to one of the following groups ( n = 8 each): the sham-operated control group, the IR group (40 min of left-lung ischaemia and 105 min of reperfusion), the IPO group (three successive cycles of 30-s reperfusion per 30-s occlusion before restoring full perfusion) and the ZnPPIX + IPO group (ZnPPIX, an inhibitor of HO-1, was injected intra-peritoneally at 20 mg kg −1 24 h prior to the experiment and the rest of the procedures were similar to that of the IPO group). Lung injury was assessed by arterial blood gas analysis, wet-to-dry lung weight ratio and tissue histological changes. The extent of lipid peroxidation was determined by measuring plasma levels of malondialdehyde (MDA) production. Expression of HO-1 was determined by immunohistochemistry. Results Lung IR resulted in a significant reduction of PaO 2 (data in IR, P < 0.05 vs. data in sham) and increase of lung wet-to-dry weight ratio, accompanied with increased MDA production and severe lung pathological morphological changes as well as a compensatory increase in HO-1 protein expression, as compared with sham (All P < 0.05). IPO markedly attenuated all the above pathological changes seen in the IR group and further increased HO-1 expression. Treatment with ZnPPIX abolished all the protective effects of post-conditioning. Conclusion It may be concluded that IPO protects IR-induced lung injury via induction of HO-1.

Protective effect of ischemic postconditioning on lung ischemia-reperfusion injury in rats and the role of heme oxygenase-1

To investigate the effect of ischemic postconditioning (IPO) on acute lung ischemia-reperfusion (I/R) injury and the protein expression of haeme oxygenase-1 (HO-1), a cytoprotective defense against oxidative injury. After being anesthetized with chloralhy- drate, forty-eight healthy SD rats were randomly divided into 6 groups (8 in each): sham operation group (S group); I/R group: left lung hilum was clamped for 40 minutes followed by 105 minutes of reperfusion; IPO group: left lung hilum was clamped for 40 minutes and postconditioned by 3 cycles of 30 seconds of reperfusion and 30 seconds of reocclusion; Hemin (HM)+ I/R group: hemin, an inducer of HO-1 was injected intraperitoneally at 40 micromol/kg/day for two consecutive days prior to 40 minutes clamping of left lung hilum; ZnPPIX+IPO group: zinc protoporphyrin IX, an inhibitor of HO-1 was injected intraperitoneally at 20 mg/kg 24 hours prior to 40 minutes clamping of left lung hilum; and HM+S group: HM was administered as in the HM+I/R group without inducing lung I/R. Arterial partial pressure of oxygen (PaO(2)) and malondialdehyde (MDA) content in serum were assessed. The left lung was removed for determination of wet/dry lung weight ratio and expression of HO-1 protein by immuno-histochemical technique and for light microscopic examination. The PaO(2) was significantly lower in all the experimental groups compared with sham group (90 mm Hg +/- 1 mm Hg). However, the values of PaO(2) in IPO (81 mm Hg +/- mm Hg) and HM+I/R (80 mm Hg+/- mm Hg) were higher than that in I/R (63 mm Hg +/- 9 mm Hg) and ZnPPIX+IPO (65 mm Hg +/- 8 mm Hg) groups (P less than 0.01). The protein expression of HO-1 in lung tissue was significantly increased in I/R group compared with S group (P less than 0.01). While the HO-1 protein expression was higher in IPO and HM+I/R groups as compared with I/R group (P less than 0.05, P less than 0.01). The lung wet/dry (W/D) weight ratio and MDA content in serum were significantly increased in I/R group as compared with S or HM+S groups (P less than 0.01), accompanied by severe lung tissue histological damage, which was attenuated either by IPO or by HM pretreatment (P less than 0.01, IPO or HM+I/R vs. I/R). The protective effect of IPO was abolished by ZnPPIX. Ischemic postconditioning can attenuate the lung ischemia-reperfusion injury through upregulating the protein expression of HO-1 that leads to reduced post-ischemic oxidative damage.