高铁血红素对心肌缺血／再灌注损伤的防护作用及其机制

目的：在大鼠急性心肌缺血／再灌ii（I／R）模型上,观察高铁血红素在钙激活中性蛋白酶（calpain）介导的心肌I／R损伤中的作用。并初步探讨其可能的机制。方法：64只雄性SD大鼠随机8组（n：8）：假手术组（sham组）、（I／R）组、MDL28170＋I／R组、单纯MDL28170组、高铁血红素＋I／R组、单纯高铁血红素组、锌原卟啉Ⅸ＋高铁血红素＋I／R组、单纯锌原卟啉Ⅸ组。采用大鼠离体心脏Langendorff灌流技术,心脏I／R后,测定左室发展压（LVDP）、心肌梗死面积、冠脉流出液中的乳酸脱氢酶（LDH）释放量。检测calpain、血红素氧化酶（HO）、和半胱氨酸天冬氨酸蛋白酶3（caspase3）活性。Westernblot观察心肌钙蛋白酶抑制蛋白（calpastatin）蛋白表达。结果：①心肌I／R后,calpain、caspase3活性明显增高。calpain抑制剂MDL28170可抑制I／R诱导的LDH释放量增加,增高LVDP,缩小心肌梗死面积。②与单纯I／R组相比,大鼠预先给予高铁血红素后,心脏HO-1活性增加,calpain和caspase3活性下降。同时,LDH释放量减少,LVDP明显增高,心肌梗死面积缩小。③I／R组心肌calpastatin表达量明显低于对照组,高铁血红素组大鼠calpastatin表达量增高。HO-1的抑制剂锌原卟啉Ⅸ可取消高铁血红素对calpastain表达量的影响,并取消其心肌保护作用。结论：高铁血红素预处理可通过抑制calpain的激活,减轻大鼠心肌I／R损伤,其机制可能与增加calpastatin蛋白表达有关。     

Protection by 3'-methoxypuerarin of rat hippocampal neurons against ischemia/reperfusion injury

3'-Methoxypuerarin (3'-MOP) is an isoflavone extracted from radix puerariae. The aim of this study was to investigate the role and the mechanism of 3'-MOP in the protection of hippocampal neurons against cerebral ischemia/reperfusion (I/R) injury in rats. I/R injury was induced by a modified four-vessel occlusion model. Rats were randomly divided into an I/R group, an I/R + 3'-MOP group and a control group. Histological changes in the neurons of the hippocampal CA1 region were observed with hematoxylin and eosine (H&E) staining. The apoptotic neurons in the hippocampal CA1 area were counted with the terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) staining. The results showed that compared with the I/R group, 3'-MOP increased the number of surviving neurons in the hippocampal CA1 region (P < 0.001) and markedly reduced the number of apoptotic pyramidal neurons (P < 0.001) after I/R injury. In conclusion, 3'-MOP can protect hippocampal neurons against I/R injury by inhibiting apoptosis.     

Hepatic ischemia/reperfusion injury involves functional TLR4 signaling in nonparenchymal cells

Endogenous ligands from damaged cells, so-called damage-associated molecular pattern molecules, can activate innate immunity via TLR4 signaling. Hepatic warm ischemia and reperfusion (I/R) injury and inflammation is largely TLR4 dependent. We produced TLR4 chimeric mice to assess whether the TLR4-dependent injury required TLR4 expression on liver parenchymal or nonparenchymal cells. Chimeric mice were produced by adoptive transfer of donor bone marrow cells into irradiated recipient animals using reciprocal combinations of TLR4 wild-type (WT; C3H/HeOuj) and TLR4 mutant (C3H/HeJ) mouse bone marrow. Wild-type chimeric mice bearing TLR4 mutant hemopoietic cells and TLR4 mutant mice transplanted with their own bone marrow-derived cells were protected from hepatic I/R and exhibited decreased JNK and NF-kappaB activation compared with WT chimeric mice transplanted with their own bone marrow. In contrast, TLR4 mutant mice transplanted with TLR4 WT bone marrow were not protected from liver I/R and demonstrated pronounced increases in JNK and NF-kappaB activation when compared with autochthonous transplanted mutant mice. In addition, depletion of phagocytes taking up gadolinium chloride failed to provide any additional protection to TLR4 mutant mice, but substantially reduced damage in WT mice after hepatic I/R. Together, these results demonstrate that TLR4 engagement on actively phagocytic nonparenchymal cells such as Kupffer cells is required for warm I/R-induced injury and inflammation in the liver.     

WISP1 mediates lung injury following hepatic ischemia reperfusion dependent on TLR4 in mice

Background

Hepatic ischemia-reperfusion injury (IRI) is a common pathological phenomenon, which causes hepatic injury as well as remote organ injuries such as the lung. Several mediators, such as oxidative stress, Ca²⁺ overload and neutrophil infiltration, have been implied in the pathogenesis of liver and remote organ injuries following reperfusion. WNT1 inducible signaling pathway protein 1 (WISP1) is an extracellular matrix protein that has been associated with the onset of several malignant diseases. Previous work in our group has demonstrated WISP1 is upregulated and contributes to proinflammatory cascades in hepatic IRI. However, the role of WISP1 in the pathogenesis of lung injury after hepatic IRI still remains unknown.

Methods

Male C57BL/6 mice were used to examine the expression and role of WISP1 in the pathogenesis of lung injuries after hepatic IRI and explore its potential mechanisms in mediating lung injuries.

Results

We found WISP1 was upregulated in lung tissues following hepatic IRI. Treatment with anti-WISP1 antibody ameliorated lung injuries with alteration of cytokine profiles. Administration with rWISP1 aggravated lung injuries following hepatic IRI through excessive production of proinflammatory cytokines and inhibition of anti-inflammatory cytokines.

Conclusions

In this study, we concluded that WISP1 contributed to lung injuries following hepatic IRI through TLR4 pathway.

     

Protective effect of nicorandil against myocardial ischemia/reperfusion injury mediated via IL33/ST2 signaling pathway

Molecular and Cellular Biochemistry - Myocardial ischemia–reperfusion injury (MI/RI), a complication of myocardial injury, is associated with high rates of mortality and disability. We aimed...     

Uncoupling protein 3 (UCP3) stimulates glucose uptake in muscle cells through a phosphoinositide 3-kinase-dependent mechanism

UCP3 is a mitochondrial membrane protein expressed in humans selectively in skeletal muscle. To determine the mechanisms by which UCP3 plays a role in regulating glucose metabolism, we expressed human UCP3 in L6 myotubes by adenovirus-mediated gene transfer and in H(9)C(2) cardiomyoblasts by stable transfection with a tetracycline-repressible UCP3 construct. Expression of UCP3 in L6 myotubes increased 2-deoxyglucose uptake 2-fold and cell surface GLUT4 2.3-fold, thereby reaching maximally insulin-stimulated levels in control myotubes. Wortmannin, LY 294002, or the tyrosine kinase inhibitor genistein abolished the effect of UCP3 on glucose uptake, and wortmannin inhibited UCP3-induced GLUT4 cell surface recruitment. UCP3 overexpression increased phosphotyrosine-associated phosphoinositide 3-kinase (PI3K) activity 2.2-fold compared with control cells (p < 0.05). UCP3 overexpression increased lactate release 1.5- to 2-fold above control cells, indicating increased glucose metabolism. In H(9)C(2) cardiomyoblasts stably transfected with UCP3 under control of a tetracycline-repressible promotor, removal of doxycycline resulted in detectable levels of UCP3 at 12 h and 2.2-fold induction at 7 days compared with 12 h. In parallel, glucose transport increased 1.3- and 2-fold at 12 h and 7 days, respectively, and the stimulation was inhibited by wortmannin or genistein. p85 association with membranes was increased 5.5-fold and phosphotyrosine-associated PI3K activity 3.8-fold. In contrast, overexpression of UCP3 in 3T3-L1 adipocytes did not alter glucose uptake, suggesting tissue-specific effects of human UCP3. Thus, UCP3 stimulates glucose transport and GLUT4 translocation to the cell surface in cardiac and skeletal muscle cells by activating a PI3K dependent pathway.     

Toll-like receptor 3 plays a role in myocardial infarction and ischemia/reperfusion injury

Innate immune and inflammatory responses mediated by Toll like receptors (TLRs) have been implicated in myocardial ischemia/reperfusion (I/R) injury. This study examined the role of TLR3 in myocardial injury induced by two models, namely, myocardial infarction (MI) and I/R. First, we examined the role of TLR3 in MI. TLR3 deficient (TLR3^−/−) and wild type (WT) mice were subjected to MI induced by permanent ligation of the left anterior descending (LAD) coronary artery for 21 days. Cardiac function was measured by echocardiography. Next, we examined whether TLR3 contributes to myocardial I/R injury. TLR3^−/− and WT mice were subjected to myocardial ischemia (45 min) followed by reperfusion for up to 3 days. Cardiac function and myocardial infarct size were examined. We also examined the effect of TLR3 deficiency on I/R-induced myocardial apoptosis and inflammatory cytokine production. TLR3^−/− mice showed significant attenuation of cardiac dysfunction after MI or I/R. Myocardial infarct size and myocardial apoptosis induced by I/R injury were significantly attenuated in TLR3^−/− mice. TLR3 deficiency increases B-cell lymphoma 2 (BCL2) levels and attenuates I/R-increased Fas, Fas ligand or CD95L (FasL), Fas-Associated protein with Death Domain (FADD), Bax and Bak levels in the myocardium. TLR3 deficiency also attenuates I/R-induced myocardial nuclear factor KappaB (NF-κB) binding activity, Tumor necrosis factor alpha (TNF-α) and Interleukin-1 beta (IL-1β) production as well as I/R-induced infiltration of neutrophils and macrophages into the myocardium. TLR3 plays an important role in myocardial injury induced by MI or I/R. The mechanisms involve activation of apoptotic signaling and NF-κB binding activity. Modulation of TLR3 may be an effective approach for ameliorating heart injury in heart attack patients.     

Pyrrolidinyl caffeamide against ischemia/reperfusion injury in cardiomyocytes through AMPK/AKT pathways

Background

Coronary heart disease is a leading cause of death in the world and therapy to reduce injury is still needed. The uncoupling of glycolysis and glucose oxidation induces lactate accumulation during myocardial ischemia/reperfusion (I/R) injury. Cell death occurs and finally leads to myocardial infarction. Caffeic acid, one of the major phenolic constituents in nature, acts as an antioxidant. Pyrrolidinyl caffeamide (PLCA), a new derivative of caffeic acid, was synthesized by our team. We aimed to investigate the effect of PLCA on hypoxia/reoxygenation (H/R) in neonatal rat ventricular myocytes (NRVM) and on myocardial I/R in rats.

Results

Cardiomyocytes were isolated and subjected to 6 h hypoxia followed by 18 h reperfusion. PLCA (0.1 to 3 μM) and metformin (30 μM) were added before hypoxia was initiated. PLCA at 1 μM and metformin at 30 μM exerted similar effects on the improvement of cell viability and the alleviation of cell apoptosis in NRVM after H/R. PLCA promoted p-AMPK, p-AKT, and GLUT4 upregulation to induce a cardioprotective effect in both cell and animal model. The accumulation of cardiac lactate was attenuated by PLCA during myocardial I/R, and infarct size was smaller in rats treated with PLCA (1 mg/kg) than in those treated with caffeic acid (1 mg/kg).

Conclusions

AMPK and AKT are synergistically activated by PLCA, which lead facilities glucose utilization, thereby attenuating lactate accumulation and cell death. The cardioprotective dose of PLCA was lower than those of metformin and caffeic acid. We provide a new insight into this potential drug for the treatment of myocardial I/R injury.     

Protection by pyruvate against glutamate neurotoxicity is mediated by astrocytes through a glutathione-dependent mechanism

Pyruvate, an endogenous metabolite of glycolysis, is an anti-toxicity agent. Recent studies have suggested possible roles for pyruvate in protecting CNS neurons from excitotoxic and metabolic insults. Utilizing cultures derived from embryonic rat cortex, the studies presented in this paper indicate that an astroglia-mediated mechanism is involved in the neuroprotective effects of pyruvate against glutamate toxicity. Glutamate-induced toxicity could be reversed by pyruvate in a mixed culture of cortex cells. Importantly, in pure neuronal cultures from the same tissue, pyruvate failed to protect against glutamate toxicity. Addition of astroglia to the pure neuronal cultures restores the ability of pyruvate to protect neurons from glutamate-induced toxicity. Our results further suggest that pyruvate can induce glia to up-regulate the synthesis of glutathione (GSH), an antioxidant that protects cells from toxins such as free radicals. Taken together, our data suggest that astroglia in mixed cultures are essential for mediating the effects of pyruvate, revealing a novel mechanism by which pyruvate, an important intermediate of tricarboxylic acid cycle in the body, may act to protect neurons from damage during insults such as brain ischemia.     

激动乙醛脱氢酶2对抗糖尿病大鼠心肌缺血/再灌注损伤的作用

目的:探讨激动乙醛脱氢酶2(ALDH2)在糖尿病大鼠心肌损伤中的作用。方法:腹腔注射55 mg/kg链脲佐菌素复制糖尿病大鼠模型,分为糖尿病组和乙醇+糖尿病组(n=8)。8周后行离体心肌缺血/再灌注(I/R),测定心室动力学指标和复灌期间冠脉流出液中乳酸脱氢酶(LDH)含量。测定空腹血糖、糖化血红蛋白(HbA1c)水平。RT-PCR和Western blot测定左心室前壁心尖组织线粒体ALDH2 mRNA和蛋白表达。结果:与正常大鼠心肌I/R相比,糖尿病大鼠左室发展压、左心室最大上升和下降速率、左室做功进一步下降,左室舒张末压抬高,复灌期冠脉流出液中LDH释放增多,心室ALDH2 mRNA和蛋白表达降低;与糖尿病大鼠心肌I/R相比,ALDH2激动剂乙醇明显促进左室发展压、左心室最大上升和下降速率、左室做功的恢复,降低左室舒张末压,同时降低HbA1c水平和LDH的释放,ALDH2 mRNA和蛋白表达增高。结论:糖尿病大鼠心肌缺血/再灌注时,心肌ALDH2表达降低;增强ALDH2在糖尿病大鼠心肌中的表达可发挥保护作用。     

SDF-1/CXCR4 mediates acute protection of cardiac function through myocardial STAT3 signaling following global ischemia/reperfusion injury

Stromal cell-derived factor-1α (SDF-1) has been reported to mediate cardioprotection through the mobilization of stem cells into injured tissue and an increase in local angiogenesis after myocardial infarction. However, little is known regarding whether SDF-1 induces acute protection following global myocardial ischemia/reperfusion (I/R) injury and if so, by what molecular mechanism. SDF-1 binding to its cognate receptor CXCR4 has been shown to activate STAT3 in a variety of cells. STAT3 is a cardioprotective factor and may mediate SDF-1/CXCR4-induced acute protection. We hypothesized that SDF-1 would improve myocardial function through CXCR4-increased STAT3 activation following acute I/R. Isolated mouse hearts were subjected to 25-min global ischemia/40-min reperfusion and divided into groups of 1) vehicle; 2) SDF-1; 3) AMD3100, a CXCR4 inhibitor; 4) SDF-1 + AMD3100; 5) Stattic, a STAT3 inhibitor; 6) SDF-1 + Stattic; 7) cardiomyocyte-restricted ablation of STAT3 (STAT3KO); 8) STAT3KO + SDF-1; 9) Ly294002, an inhibitor of the Akt pathway; and 10) SDF-1 + Ly294002. Reagents were infused into hearts within 5 min before ischemia. SDF-1 administration significantly improved postischemic myocardial functional recovery in a dose-dependent manner. Additionally, pretreatment with SDF-1 reduced cardiac apoptotic signaling and increased myocardial STAT3 activation following acute I/R. Inhibition of the SDF-1 receptor CXCR4 neutralized these protective effects by SDF-1 in hearts subjected to I/R. Notably, inhibition of the STAT3 pathway or use of STAT3KO hearts abolished SDF-1-induced acute protection following myocardial I/R. Our results represent the first evidence that the SDF-1/CXCR4 axis upregualtes myocardial STAT3 activation and, thereby, mediates acute cardioprotection in response to global I/R.     

Overexpression of HSPA12B protects against cerebral ischemia/reperfusion injury via a PI3K/Akt-dependent mechanism

Background and purpose: HSPA12B is a newly discovered member of the Hsp70 family proteins. This study investigated the effects of HSPA12B on focal cerebral ischemia/reperfusion (I/R) injury in mice. Methods: Transgenic mice overexpressing human HSPA12B (Tg) and wild-type littermates (WT) were subjected to 60 min of middle cerebral artery occlusion to induce ischemia and followed by reperfusion (I/R). Neurological deficits, infarct volumes and neuronal death were examined at 6 and 24 hrs after reperfusion. Blood–brain-barrier (BBB) integrity and activated cellular signaling were examined at 3 hrs after reperfusion. Results: After cerebral I/R, Tg mice exhibited improvement in neurological deficits and decrease in infarct volumes, when compared with WT I/R mice. BBB integrity was significantly preserved in Tg mice following cerebral I/R. Tg mice also showed significant decreases in cell injury and apoptosis in the ischemic hemispheres. We observed that overexpression of HSPA12B activated PI3K/Akt signaling and suppressed JNK and p38 activation following cerebral I/R. Importantly, pharmacological inhibition of PI3K/Akt signaling abrogated the protection against cerebral I/R injury in Tg mice. Conclusions: The results demonstrate that HSPA12B protects the brains from focal cerebral I/R injury. The protective effect of HSPA12B is mediated though a PI3K/Akt-dependent mechanism. Our results suggest that HSPA12B may have a therapeutic potential against ischemic stroke.     

Response to myocardial ischemia/reperfusion injury involves Bnip3 and autophagy

Ischemia and reperfusion (I/R) injury is associated with extensive loss of cardiac myocytes. Bnip3 is a mitochondrial pro-apoptotic Bcl-2 protein which is expressed in the adult myocardium. To investigate if Bnip3 plays a role in I/R injury, we generated a TAT-fusion protein encoding the carboxyl terminal transmembrane deletion mutant of Bnip3 (TAT-Bnip3DeltaTM) which has been shown to act as a dominant negative to block Bnip3-induced cell death. Perfusion with TAT-Bnip3DeltaTM conferred protection against I/R injury, improved cardiac function, and protected mitochondrial integrity. Moreover, Bnip3 induced extensive fragmentation of the mitochondrial network and increased autophagy in HL-1 myocytes. 3D rendering of confocal images revealed fragmented mitochondria inside autophagosomes. Enhancement of autophagy by ATG5 protected against Bnip3-mediated cell death, whereas inhibition of autophagy by ATG5K130R enhanced cell death. These results suggest that Bnip3 contributes to I/R injury which triggers a protective stress response with upregulation of autophagy and removal of damaged mitochondria.     

N-乙酰-L-半胱氨酸减轻大鼠胃缺血/再灌注损伤的机制

为研究N-乙酰-L-半胱氨酸(N-acetylcysteine,NAC)减轻大鼠胃缺血/再灌注(gastric ischemia/reperfusion,GI/R)损伤的机制,在大鼠股静脉注射NAC(150mg/kg),夹闭大鼠腹腔动脉30min,再灌注1h制备GI/R模型。取胃后计数胃黏膜损伤指数(gastric mucosal damage index,GMDI),用原位检测(TUNEL)法观察胃黏膜细胞的凋亡,用免疫印迹法测定胃黏膜组织中p-ERK,p-JNK和NF-κB的表达,用RT-PCR法检测TNF-α,Caspase-3的mRNA表达。结果显示,NAC可以减轻GI/R损伤大鼠胃黏膜细胞的凋亡;促进大鼠I/R损伤的胃黏膜组织中p-ERK蛋白的表达,抑制p-JNK和NF-κB的蛋白表达,同时也抑制TNF-α mRNA和Caspase-3 mRNA的表达。股静脉给予辣椒素受体(vanilloid receptor subtype1,VR1)拮抗剂(capsaizepin,CPZ)400mg/kg,能逆转NAC对大鼠GI/R损伤的保护作用。以上结果提示:NAC对大鼠GI/R损伤具有减轻作用,其保护机制可能是通过上调胃黏膜组织中p-ERK,下调p-JNK和NF-κB实现的,且这种保护作用可能与VR1有关。     

雷米普利与BQ-123合用对大鼠在体心肌缺血／再灌注损伤的保护作用

目的：研究雷米普利与BQ-123合用对大鼠在体心肌缺血／再灌注损伤的影响。方法：健康雄性Wistar大鼠随机分为5组,制备缺血30min再灌注120min模型,采用雷米普利、BQ-123单用及两药联合应用的方式处理实验动物。观察两药合用对大鼠在体心肌缺血／再灌注损伤的保护作用。观察动物心率、血压、心电图ST-段变化,记录缺血期室性心律失常;检测血浆CK及LDH活力;心肌HE染色和TIC染色,定性和定量检测心肌梗死情况。结果：与I／R组比较,各给药组ST-段均明显降低;缺血期室性心律失常（VA）出现时间明显推迟且持续时间明显缩短,联合给药组作用更为显著;心律失常发生率明显降低;血浆CK及LDH活力显著降低且联合给药组降低更为显著;梗死面积明显缩小,心肌损伤程度明显减轻,其中联合给药组变化更为显著。结论：雷米普利、BQ-123单用及联合应用均对大鼠在体心肌缺血／再灌注损伤有保护作用,且两药合用在推迟缺血期VA的出现时间,缩短其持续时间,减少CK及LDH漏出,缩小心肌梗死面积方面优于两药各自单用。     

Long-term aerobic exercise protects the heart against ischemia/reperfusion injury via PI3 kinase-dependent and Akt-mediated mechanism

Objective Physical activity has been shown to improve cardiovascular function and to be beneficial to type 2 diabetic patients. However, the effects of aerobic exercise (AE) on myocardial ischemia/reperfusion (MI/R) are largely unclear. Therefore, the aims of the present study were to determine whether long-term AE can protect the heart against I/R injury, and if so, to investigate the underlying mechanism. Methods Adult male Sprague–Dawley rats were randomly subjected to 8 weeks of either sedentary or free-loading swimming exercise (3 h/day, 5 d/week). Then the animals were subjected to 30 min MI followed by 4 h R. Arterial blood pressure and left ventricular pressure (LVP) were monitored throughout the whole MI/R procedure. Plasma creatine kinase (CK) and lactate dehydrogenase (LDH) activities were measured spectrophotometrically. Myocardial infarction and myocardial apoptosis (TUNEL analysis) were determined in a blinded manner. Results MI/R caused significant cardiac dysfunction and myocardial apoptosis (strong TUNEL-positive staining). Compared with sedentary group, rats subjected to 8 weeks of AE showed protection against MI/R as evidenced by reduced myocardial infarction (26.8 ± 1.5% vs. 35.3 ± 2.4%, n = 8, P < 0.05), inhibited cardiomyocyte apoptosis (decreased apoptotic index (12.4 ± 1.1% vs. 21.0 ± 1.7%, n = 8, P < 0.01) and decreased myocardial caspase-3 activity), decreased plasma CK and LDH activities and improved recovery of cardiac systolic/diastolic function (including LVSP and ±LVdP/dt) at the end of R. Moreover, exercise resulted in 1.7-fold, 2.5-fold and 2.5-fold increases in Akt expression, Akt phosphorylation and glycogen synthase kinase-3β phosphorylation in I/R myocardium, respectively (n = 3, all P < 0.05). More importantly, treatment with wortmannin, a PI3 kinase inhibitor, 15 min before R not only significantly blocked Akt phosphorylation (P < 0.05) in exercise rats, but also abolished long-term AE-induced cardioprotection for the I/R heart as manifested by increased apoptosis and myocardial infarction, and reduced cardiac function. Conclusion Long-term AE exerts cardioprotective effect against MI/R injury, including anti-cardiomyocyte apoptosis, which is at least partly via PI3 kinase-dependent and Akt-mediated mechanism.     

CpG-ODN,the TLR9 agonist,attenuates myocardial ischemia/reperfusion injury: Involving activation of PI3K/Akt signaling

BackgroundToll-like receptors (TLRs) have been implicated in myocardial ischemia/reperfusion (I/R) injury. The TLR9 ligand, CpG-ODN has been reported to improve cell survival. We examined effect of CpG-ODN on myocardial I/R injury.MethodsMale C57BL/6 mice were treated with either CpG-ODN, control-ODN, or inhibitory CpG-ODN (iCpG-ODN) 1 h prior to myocardial ischemia (60 min) followed by reperfusion. Untreated mice served as I/R control (n = 10/each group). Infarct size was determined by TTC straining. Cardiac function was examined by echocardiography before and after myocardial I/R up to 14 days.ResultsCpG-ODN administration significantly decreased infarct size by 31.4% and improved cardiac function after myocardial I/R up to 14 days. Neither control-ODN nor iCpG-ODN altered I/R-induced myocardial infarction and cardiac dysfunction. CpG-ODN attenuated I/R-induced myocardial apoptosis and prevented I/R-induced decrease in Bcl2 and increase in Bax levels in the myocardium. CpG-ODN increased Akt and GSK-3β phosphorylation in the myocardium. In vitro data suggested that CpG-ODN treatment induced TLR9 tyrosine phosphorylation and promoted an association between TLR9 and the p85 subunit of PI3K. Importantly, PI3K/Akt inhibition and Akt kinase deficiency abolished CpG-ODN-induced cardioprotection.ConclusionCpG-ODN, the TLR9 ligand, induces protection against myocardial I/R injury. The mechanisms involve activation of the PI3K/Akt signaling pathway.