2-4-diaminohypoxanthine and Myocardial-Ischemia

Previously we showed that Brown Norway (BN/Mcw) rats are more resistant to myocardial ischemia-reperfusion (I/R) injury than Dahl S (SS/Mcw) rats due to increased nitric oxide (x NO) generation secondary to increased heat shock protein 90 (HSP90) association with endothelial nitric oxide synthase (NOS3). Here we determined whether increased resistance to I/R injury in BN/Mcw hearts is also related to tetrahydrobiopterin (BH(4)) and GTP cyclohydrolase I (GCH-1), the rate-limiting enzyme for BH(4) synthesis. We observed that BH(4) supplementation via sepiapterin (SP) and inhibition of GCH-1 via 2,4-diamino-6-hydroxypyrimidine (DAHP) differentially modulate cardioprotection and that SP alters the association of HSP90 with NOS3. BH(4) levels were significantly higher and 7,8-dihydrobiopterin (BH(2)) levels were significantly lower in BN/Mcw than in SS/Mcw hearts. The BH(4)-to-BH(2) ratio in BN/Mcw was more than two times that in SS/Mcw hearts. After I/R, BH(4) decreased and BH(2) increased in hearts from both strains compared with their preischemia levels. However, the increase in BH(2) in SS/Mcw hearts was significantly higher than in BN/Mcw hearts. Real-time PCR revealed that BN/Mcw hearts contained more GCH-1 transcripts than SS/Mcw hearts. SP increased recovery of left ventricular developed pressure (rLVDP) following I/R as well as decreased superoxide (O(2)(x-)) and increased x NO in SS/Mcw hearts but not in BN/Mcw hearts. DAHP decreased rLVDP as well as increased O(2)(x-) and decreased x NO in BN/Mcw hearts compared with controls but not in SS/Mcw hearts. SP increased the association of HSP90 with NOS3. These data indicate that BH(4) mediates resistance to I/R by acting as a cofactor and enhancing HSP90-NOS3 association.

Topics: Animals; Biopterins; Disease Models, Animal; Enzyme Inhibitors; Gene Expression Regulation, Enzymologic; GTP Cyclohydrolase; HSP90 Heat-Shock Proteins; Hypoxanthines; Myocardial Ischemia; Myocardial Reperfusion Injury; Myocytes, Cardiac; Nitric Oxide; Nitric Oxide Synthase Type III; Pterins; Rats; Rats, Inbred BN; Rats, Inbred Dahl; RNA, Messenger; Species Specificity; Superoxides; Ventricular Function, Left; Ventricular Pressure

It has recently been shown that nitric oxide synthase in the presence of suboptimal levels of tetrahydrobiopterin (BH(4)), an essential cofactor of nitric oxide synthase, may favor increased production of oxygen free radicals. This study was designed to define the role of BH(4) in myocardial ischemia-reperfusion injury.. Isolated perfused rat hearts were subjected to 37 degrees C ischemia and reperfusion. Hearts were received with BH(4) or vehicle for 5 min just before ischemia and during the first 5 min of the reperfusion period. The effects of BH(4) on left ventricular function, myocardial contents of lipid peroxidation and high energy phosphates, and levels of lactate dehydrogenase and nitrite plus nitrate in perfusate before ischemia and after reperfusion were estimated. Moreover, the effect of BH(4) given with 2,4-diamino-6-hydroxypyrimidine (DAHP), a selective inhibitor of BH(4) production, intraperitoneally 24 h before the experiments were estimated.. BH(4) improved contractile and metabolic abnormalities in reperfused hearts. Furthermore, BH(4) significantly alleviated ischemic contracture during ischemia, and restored diminished perfusate levels of nitrite plus nitrate after reperfusion. On the other hand, DAHP-treatment aggravated ischemia-reperfusion induced functional and metabolic abnormalities. Administration of BH(4) improved DAHP-induced functional and metabolic abnormalities.. Results demonstrated that BH(4) lessens ischemia-reperfusion injury in isolated perfused rat hearts. Conversely, deficiency of BH(4) seems to accelerate endothelial dysfunction and myocardial ischemia-reperfusion injury. Present data may be compatible with the hypothesis that nitric oxide synthase in the presence of insufficiency of BH(4) serve as the cause of oxidative injury.

Topics: Animals; Antioxidants; Biopterins; Disease Models, Animal; Endothelium, Vascular; Heart; Hypoxanthines; In Vitro Techniques; Lipid Peroxidation; Male; Myocardial Ischemia; Myocardial Reperfusion Injury; Nitric Oxide; Rats; Rats, Sprague-Dawley; Ventricular Function, Left

A deficit in the endothelial production of nitric oxide (NO) is associated with the sequelae of reperfusion injury. Because endothelial NO synthesis depends on the cofactor tetra-hydrobiopterin (BH4), we hypothesized that depletion of this cofactor underlies the reduction of endothelium-dependent dilation in reperfusion injury.. After occlusion of the left anterior descending coronary artery of a pig for 60 minutes followed by 90 minutes of reperfusion (ischemia/reperfusion), hearts were removed and the arterioles were isolated, cannulated, pressurized, and placed on an inverted microscope stage. Dose responses to the endothelium-independent dilator sodium nitroprusside and the endothelium-dependent dilators serotonin, A23187, and substance P were obtained under control conditions, after incubation with sepiapterin (intracellularly converted to BH4) or synthetic BH4 6-methyltetrahydropterin (MH4), and again after their washout. After ischemia/reperfusion, sodium nitroprusside maximally dilated arterioles (99 +/- 3%), whereas relaxation to serotonin, A23187, and substance P was significantly reduced (19 +/- 9%, 44 +/- 9%, and 54 +/- 8%, respectively). During incubation with sepiapterin (1 mumol/L) or MH4 (10 mumol/L), endothelium-dependent dilation was significantly enhanced (P < .05), whereas the response to sodium nitroprusside was unaltered. After washout, the vasodilatory responses were not significantly different from the initial ischemia/reperfusion responses. Sepiapterin and MH4 did not affect vasodilatory responses in vessels obtained from nonischemic control hearts. As after ischemia/reperfusion, incubation of control vessels with 2,4-diamino-6-hydroxypyrimidine, an inhibitor of GTP cyclohydrolase I, decreased endothelium-dependent vasodilation, which was restored in the presence of sepiapterin or MH4.. These data indicate that exogenous administration of sepiapterin or MH4 restores the response to endothelium-dependent vasodilators in pig coronary arterioles after ischemia/ reperfusion. We therefore conclude that ischemia/reperfusion alters the availability or production of BH4, which contributes to blunted endothelial nitroxidergic vasodilation.

Topics: Animals; Biopterins; Endothelium, Vascular; Enzyme Inhibitors; Hemodynamics; Hypoxanthines; In Vitro Techniques; Myocardial Ischemia; Myocardial Reperfusion Injury; Pteridines; Pterins; Reference Values; Swine; Vasodilation