PPAR-alpha activation protects the type 2 diabetic myocardium against ischemia-reperfusion injury: involvement of the PI3-Kinase/Akt and NO pathway.

Several clinical studies have shown the beneficial cardiovascular effects of fibrates in patients with diabetes and insulin resistance. The ligands of peroxisome proliferator-activated receptor-alpha (PPAR-alpha) reduce ischemia-reperfusion injury in nondiabetic animals. We hypothesized that the activation of PPAR-alpha would exert cardioprotection in type 2 diabetic Goto-Kakizaki (GK) rats, involving mechanisms related to nitric oxide (NO) production via the phosphatidylinositol 3-kinase (PI3K)/Akt pathway. GK rats and age-matched Wistar rats (n >or= 7) were given either 1) the PPAR-alpha agonist WY-14643 (WY), 2) dimethyl sulfoxide (DMSO), 3) WY and the NO synthase inhibitor N(G)-nitro-l-arginine (l-NNA), 4) l-NNA, 5) WY and the PI3K inhibitor wortmannin, or 6) wortmannin alone intravenously before a 35-min period of coronary artery occlusion followed by 2 h of reperfusion. Infarct size (IS), expression of endothelial NO synthase (eNOS), inducible NO synthase, and Akt as well as nitrite/nitrate were determined. The IS was 75 +/- 3% and 72 +/- 4% of the area at risk in the Wistar and GK DMSO groups, respectively. WY reduced IS to 56 +/- 3% in Wistar (P < 0.05) and to 46 +/- 5% in GK rats (P < 0.001). The addition of either l-NNA or wortmannin reversed the cardioprotective effect of WY in both Wistar (IS, 70 +/- 5% and 65 +/- 5%, respectively) and GK (IS, 66 +/- 4% and 64 +/- 4%, P < 0.05, respectively) rats. The expression of eNOS and eNOS Ser1177 in the ischemic myocardium from both strains was increased after WY. The expression of Akt, Akt Ser473, and Akt Thr308 was also increased in the ischemic myocardium from GK rats following WY. Myocardial nitrite/nitrate levels were reduced in GK rats (P < 0.05). The results suggest that PPAR-alpha activation protects the type 2 diabetic rat myocardium against ischemia-reperfusion injury via the activation of the PI3K/Akt and NO pathway.

Adiponectin protects against myocardial ischaemia-reperfusion injury via AMP-activated protein kinase, Akt, and nitric oxide.

AIMS: Cardiovascular disease and type 2 diabetes mellitus are associated with low plasma concentration of adiponectin. The aim of this study was to investigate whether adiponectin exerts cardioprotective effects during myocardial ischaemia-reperfusion and whether this effect is related to the production of nitric oxide (NO). METHODS AND RESULTS: Isolated rat hearts were subjected to 30 min of either global or local ischaemia followed by 60 min of reperfusion. The hearts received vehicle, adiponectin (3 microg/mL), the NO-synthase inhibitor nitro-l-arginine (L-NNA) (0.1 mM), or a combination of adiponectin and L-NNA at the onset of ischaemia. Haemodynamics, infarct size, and expression of endothelial NO-synthase (eNOS), AMP-activated protein kinase (AMPK), and Akt were determined. Adiponectin significantly increased left ventricular function and coronary flow during reperfusion in comparison with the vehicle group. Co-administration of L-NNA abrogated the improvement in myocardial function induced by adiponectin. Infarct size following local ischaemia-reperfusion was 40 +/- 6% of the area at risk in the vehicle group. Adiponectin reduced infarct size to 19 +/- 2% (P < 0.01). L-NNA did not affect infarct size per se but abolished the protective effect of adiponectin (infarct size 40 +/- 5%). Phosphorylation of eNOS Ser1177, AMPK Thr172, and Akt Ser 473 was increased in the adiponectin group (P < 0.05). CONCLUSION: Adiponectin protects from myocardial contractile dysfunction and limits infarct size following ischaemia and reperfusion by a mechanism involving activation of AMPK and production of NO.

Cardioprotective effect of rosuvastatin in vivo is dependent on inhibition of geranylgeranyl pyrophosphate and altered RhoA membrane translocation.

Hydroxymethyl glutaryl (HMG)-coenzyme A (CoA) reductase inhibitors (statins) protect the myocardium against ischemia-reperfusion injury via a mechanism unrelated to cholesterol lowering. Statins may inhibit isoprenylation and thereby prevent activation of proteins such as RhoA. We hypothesized that statins protect the myocardium against ischemia-reperfusion injury via a mechanism involving inhibition of geranylgeranyl pyrophosphate synthesis and translocation of RhoA to the plasma membrane. Sprague-Dawley rats were given either the HMG-CoA reductase inhibitor rosuvastatin, geranylgeranyl pyrophosphate dissolved in methanol, the combination of rosuvastatin and geranylgeranyl pyrophosphate, rosuvastatin and methanol, or distilled water (control) by intraperitoneal injection for 48 h before ischemia-reperfusion. Animals were anesthetized and either subjected to 30 min of coronary artery occlusion followed by 2 h of reperfusion where at infarct size was determined, or the expression of RhoA protein was determined in cytosolic and membrane fractions of nonischemic myocardium. There were no significant differences in hemodynamics between the control group and the other groups before ischemia or during ischemia and reperfusion. The infarct size was 80 +/- 3% of the area at risk in the control group. Rosuvastatin reduced infarct size to 64 +/- 2% (P<0.001 vs. control). Addition of geranylgeranyl pyrophosphate (77 +/- 2%, P<0.01 vs. rosuvastatin) but not methanol (65 +/- 2%, not significant vs. rosuvastatin) abolished the cardioprotective effect of rosuvastatin. Geranylgeranyl pyrophosphate alone did not affect infarct size per se (84 +/- 2%). Rosuvastatin increased the cytosol-to-membrane ratio of RhoA protein in the myocardium (P<0.05 vs. control). These changes were abolished by addition of geranylgeranyl pyrophosphate. We conclude that the cardioprotection and the increase of the RhoA cytosol-to-membrane ratio induced by rosuvastatin in vivo are blocked by geranylgeranyl pyrophosphate. The inhibition of geranylgeranyl pyrophosphate formation and subsequent modulation of cytosol/membrane-bound RhoA are of importance for the protective effect of statins against myocardial ischemia-reperfusion injury.

Cardioprotection mediated by rosiglitazone, a peroxisome proliferator-activated receptor gamma ligand, in relation to nitric oxide.

UNLABELLED: Activation of peroxisome proliferator-activated receptor (PPAR) gamma protects from myocardial ischemia/reperfusion injury. The aim of the study was to investigate whether the cardioprotective effect of PPARgamma is related to nitric oxide (NO). METHODS: Wild type (WT) and endothelial NO synthase (eNOS) knockout (KO) mice received 3 mg/kg of the PPARgamma agonist rosiglitazone or vehicle (n = 6-9 in each group) i. p. 45 min before anesthesia. The hearts were isolated, perfused in a Langendorff mode and subjected to global ischemia and 30 min reperfusion. The hearts of another two groups of WT mice received the NOS inhibitor L-NNA (100 micromol/l) or vehicle in addition to pre-treatment with vehicle or rosiglitazone. RESULTS: In the WT heart, rosiglitazone increased the recovery of left ventricular function and coronary flow following ischemia in comparison with the vehicle group.L-NNA did not affect recovery per se but significantly blunted the improvement in the recovery of left ventricular function induced by rosiglitazone. In the KO group rosiglitazone suppressed the recovery of myocardial function following ischemia. Expression of eNOS was not affected, but phosphorylated eNOS was significantly increased by rosiglitazone in the WT hearts (P < 0.05). CONCLUSION: These results suggest that the cardioprotective effect of the PPARgamma agonist rosiglitazone is mediated via NO by phosphorylation of eNOS.

Endothelin-1 exerts a preconditioning-like cardioprotective effect against ischaemia/reperfusion injury via the ET(A) receptor and the mitochondrial K(ATP) channel in the rat in vivo.

In vitro studies have demonstrated that endothelin-1 (ET-1) given before myocardial ischaemia may evoke a preconditioning (PC)-like cardioprotective effect. The first aim of this study was to investigate whether administration of ET-1 before ischaemia exerts cardioprotection against ischaemia/reperfusion injury in vivo and to determine involvement of the ET-1 receptor subtype. The second aim was to examine the role of mitochondrial ATP-sensitive K+ channels (mitoK(ATP)) as a mediator of this cardioprotection. Anaesthetised open-chest Wistar rats were subjected to 30 min of coronary artery occlusion followed by 2 h reperfusion (I/R). In protocol I, the first group was subjected to I/R only (control, n=10). In the second (n=10) group, PC was elicited by three 5 min cycles of coronary artery occlusion, separated by 5 min reperfusion before I/R. The third (n=6) and fourth (n=7) groups were given ET-1 intravenous (i.v.) during three 5 min infusion periods separated by 5 min before I/R. The fourth group was in addition given the ET(A) receptor antagonist LU 135252 5 min before the infusions of ET-1. In protocol II, the first group was I/R control as in protocol I (n=8). The second (n=6), third (n=7) and fourth (n=7) groups were given ET-1 as in protocol I. The third group was in addition given the nonselective K(ATP) channel antagonist glibenclamide (Glib) 30 min before the ET-1 infusions and the fourth group the selective mitoK(ATP) channel antagonist 5-hydroxydecanoic acid (5-HD) 5 min before I/R. There were no significant differences in MAP or heart rate between the groups during I/R. In protocol I, PC reduced IS compared to the control group (10+/-3 vs 35+/-5%, P<0.01). Infusion of ET-1 also reduced IS (to 14+/-3%, P<0.05 vs control). The ET(A) receptor antagonist blocked the reduction in IS induced by ET-1 (IS 47+/-8% after LU+ET-1; P< 0.05 vs ET-1). In protocol II, Glib and 5-HD abolished the cardioprotective effect induced by ET-1 (IS 48+/-7% after Glib+ET-1 and 42+/-5% after ET-1+5-HD vs 18+/-4% after ET-1 alone; P<0.05). In conclusion, administration of ET-1 before ischaemia resulted in a PC-like cardioprotective effect. This effect is mediated via the ET(A) receptor and activation of mitoK(ATP) channels.

Cardioprotective effect induced by brief exposure to nitric oxide before myocardial ischemia-reperfusion in vivo.

Administration of nitric oxide (NO) donors during ischemia and reperfusion protects from myocardial injury. However, whether administration of an NO donor during a brief period prior to ischemia protects the myocardium and the endothelium against ischemia-reperfusion injury in vivo is unknown. To study this possibility anesthetized pigs were subjected to 45-min ligation of the left anterior descending coronary artery (LAD) followed by 4h of reperfusion. In initial dose-finding experiments, vehicle or three different doses of the NO donor S-nitroso-N-acetyl-D,L-penicillamin (SNAP; 0.1; 0.5; 2.5 micromol) were infused into the LAD for 3 min starting 13 min during ischemia. Only the 0.5 micromol dose of SNAP reduced infarct size (from 85+/-3% of the area at risk in the vehicle group to 63+/-3% in the SNAP-treated group; p<0.01). There were no significant differences in hemodynamics in the vehicle and SNAP groups during ischemia-reperfusion. Endothelium-dependent dilatation of coronary microvasculature induced by substance P was larger in the SNAP group than in the vehicle group. Myeloperoxidase activity was lower in the ischemic/reperfused myocardial area of pigs given SNAP (4.97+/-0.61 U/g) than in vehicle-treated pigs (8.45+/-0.25 U/g; p<0.05). It is concluded that intracoronary administration of the NO donor SNAP for a brief period before ischemia reduces infarct size, attenuates neutrophil accumulation, and improves endothelial function. These results suggest that NO exerts a classic preconditioning-like protection against ischemia-reperfusion injury in vivo in a narrow concentration range.