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Significant advances in our understanding of the ability of nitric oxide synthases (NOS) to modulate cardiac function have provided key insights into the role NOS play in the regulation of excitation–contraction (EC) coupling in health and disease. Through both cGMP-dependent and cGMP-independent (e.g. S-nitrosylation) mechanisms, NOS have the ability to alter intracellular Ca2 + handling and the myofilament response to Ca2 +, thereby impacting the systolic and diastolic performance of the myocardium. Findings from experiments using nitric oxide (NO) donors and NOS inhibition or gene deletion clearly implicate dysfunctional NOS as a critical contributor to many cardiovascular disease states. However, studies to date have only partially addressed NOS isoform-specific effects and, more importantly, how subcellular localization of NOS influences ion channels involved in myocardial EC coupling and excitability. In this review, we focus on the contribution of each NOS isoform to cardiac dysfunction and on the role of uncoupled NOS activity in common cardiac disease states, including heart failure, diabetic cardiomyopathy, ischemia/reperfusion injury and atrial fibrillation. We also review evidence that clearly indicates the importance of NO in cardioprotection. This article is part of a Special Issue entitled "Redox Signalling in the Cardiovascular System".

Myocardial constitutive No production depends on the activity of both endothelial and neuronal NOS (eNOS and nNOS, respectively). Stimulation of myocardial β(3)-adrenergic receptor (β(3)-AR) produces a negative inotropic effect that is dependent on eNOS. We evaluated whether nNOS also plays a role in β(3)-AR signaling and found that the β(3)-AR-mediated reduction in cell shortening and [Ca(2+)](i) transient amplitude was abolished both in eNOS(-/-) and nNOS(-/-) left ventricular (LV) myocytes and in wild type LV myocytes after nNOS inhibition with S-methyl-l-thiocitrulline. LV superoxide (O(2)()) production was increased in nNOS(-/-) mice and reduced by l-N(ω)-nitroarginine methyl ester (l-NAME), indicating uncoupling of eNOS activity. eNOS S-glutathionylation and Ser-1177 phosphorylation were significantly increased in nNOS(-/-) myocytes, whereas myocardial tetrahydrobiopterin, eNOS Thr-495 phosphorylation, and arginase activity did not differ between genotypes. Although inhibitors of xanthine oxidoreductase (XOR) or NOX2 NADPH oxidase caused a similar reduction in myocardial O(2)(), only XOR inhibition reduced eNOS S-glutathionylation and Ser-1177 phosphorylation and restored both eNOS coupled activity and the negative inotropic and [Ca(2+)](i) transient response to β(3)-AR stimulation in nNOS(-/-) mice. In summary, our data show that increased O(2)() production by XOR selectively uncouples eNOS activity and abolishes the negative inotropic effect of β(3)-AR stimulation in nNOS(-/-) myocytes. These findings provide unequivocal evidence of a functional interaction between the myocardial constitutive NOS isoforms and indicate that aspects of the myocardial phenotype of nNOS(-/-) mice result from disruption of eNOS signaling.

Resumen La homocisteina (HCY) es un aminoacido cuya elevacion sanguinea se asocia con el desarrollo de enfermedades de tipo vascular, neurologico y reproductivo. Sus niveles plasmaticos varian en funcion de la raza, el sexo, la edad y otros factores ambientales. En la presente revision se aborda su metabolismo, su fisiopatologia y las consecuencias clinicas de su elevacion. Un enfasis especial se presta al empleo del raton como modelo experimental en este campo, ya que su uso ha puesto de manifiesto la importancia de la dieta en la regulacion de la HCY. Esto, unido al desarrollo de animales modificados geneticamente con hiperhomocisteinemia, esta permitiendo una rapida caracterizacion de los mecanismos moleculares implicados en la accion in vivo de la HCY. Ademas, la combinacion de estos modelos con otros modificados geneticamente permite definir la influencia de la combinacion de factores de riesgo en el desarrollo de diversas patologias. A su vez, la exploracion en estos nuevos modelos de factores ambientales y/o farmacologicos contribuye de este modo a explicar muchas de las evidencias epidemiologicas en humanos asi como el tratamiento mas adecuado para cada condicion.

Folic acid is a vitamin that when used as a dietary supplementation can improve endothelial function. To assess the effect of folic acid on the development of atherosclerosis, male apolipoprotein E-deficient mice fed a standard chow diet received either water (control group) or an aqueous solution of folic acid that provided a dose of 75 microg/kg/day, for ten weeks. At the time of sacrifice, blood was drawn and the heart removed. The study measured plasma homocysteine, lipids, lipoproteins, low-density lipoprotein (LDL) oxidation, isoprostane, paraoxonase, and apolipoproteins, and aortic atherosclerotic areas. In folic acid-treated animals, total cholesterol, mainly carried in very low-density and low-density lipoproteins, increased significantly, and homocysteine, HDL cholesterol, paraoxonase, and triglyceride levels did not change significantly. Plasma isoprostane and apolipoprotein (apo) B levels decreased. The resistance of LDL to oxidization and plasma apoA-I and apoA-IV levels increased with a concomitant decrease in the area of atherosclerotic lesions. The administration of folic acid decreased atherosclerotic lesions independently of plasma homocysteine and cholesterol levels, but was associated with plasma levels of apolipoproteins A-I, A-IV and B, and decreased oxidative stress.

Trans-10, cis-12-conjugated linoleic acid (CLA)-enriched diets promote atherosclerosis in mice despite increasing blood concentrations of HDL cholesterol. This suggests that under these conditions, the HDL apolipoproteins (apo) produced are abnormal. To test this hypothesis, apoE-deficient mice were fed a Western-type diet enriched with linoleic acid (control), cis-9, trans-11-CLA or trans-10, cis-12-CLA (1.0% wt/wt) for 12 wk, and the effects on HDL metabolism and apoC-III levels recorded. Compared with the control and cis-9, trans-11-CLA mice, those fed the trans-10, cis-12-CLA diet had significantly higher HDL cholesterol concentrations, and had a higher incidence of hypertriglyceridemia and hepatic steatosis. Plasma apoA-I and paraoxonase concentrations were significantly lower in the trans-10, cis-12-CLA group than in the cis-9, trans-11-CLA group. These reductions were associated with decreased hepatic expression of these proteins and a shift toward lipid-poor apolipoprotein particles. The plasma apoA-II concentration increased with its corresponding mRNA concentration in the liver, and was preferentially bound to HDL in the trans-10, cis-12-CLA mice, thus explaining the increased HDL cholesterol concentrations in this group. Significant, positive associations were found between apoA-II and C-III (r=0.883, P

Apolipoprotein A-IV is a member of the apo A-I/C-III/A-IV gene cluster. In order to investigate its hypothetical coordinated regulation, an acute phase was induced in pigs by turpentine oil injection. The hepatic expression of the gene cluster as well as the plasma levels of apolipoproteins were monitored at different time periods. Furthermore, the involvement of the inflammatory mediators’ interleukins 1 and 6 and tumor necrosis factor in the regulation of this gene cluster was tested in cultured pig hepatocytes, incubated with those mediators and apo A-I/C-III/A-IV gene cluster expression at the mRNA level was measured. In response to turpentine oil-induced inflammation, a decreased hepatic apo A-IV mRNA expression was observed (independent of apo A-I and apo C-III mRNA) not correlating with the plasma protein levels. The distribution of plasma apo A-IV experienced a shift from HDL to larger particles. In contrast, the changes in apo A-I and apo C-III mRNA were reflected in their corresponding plasma levels. Addition of cytokines to cultured pig hepatocytes also decreased apo A-IV and apo A-I mRNA levels. All these results show that the down-regulation of apolipoprotein A-I and A-IV messages in the liver may be mediated by interleukin 6 and TNF-a. The well-known HDL decrease found in many different acute-phase responses also appears in the pig due to the decreased expression of apolipoprotein A-I and the enlargement of the apolipoprotein A-IV-containing HDL. 2005 Elsevier Ltd. All rights reserved.

La presente revision aborda el metabolismo lipoproteico comparado y la induccion de la aterosclerosis con sus controversias en varios modelos animales pertenecientes a un amplio espectro evolutivo que abarca desde los roedores (raton, conejo, rata, hamster, cobaya), las aves (paloma), los cetartiodactilos (cerdo) y los carnivoros (perro) hasta los primates (macacos, Rhesus, mono verde africano).

Rationale: Neuronal nitric oxide synthase (nNOS) expression and activity in atria are markedly reduced from patients and goats with atrial fibrillation (AF). Whether loss of nNOS contributes to AF-induced atrial electrical remodelling and its upstream mechanism remain unclear.Methodology: Whole-cell patch clamp was used to record action potentials (APs) and ion currents. N/n: number of patients or mice/ number of myocytes.Results: Inhibition of nNOS by S-methylthiocitrulline (SMTC, N/n: 12/45) induces a significant reduction in AP duration (APD) and APD rate-dependent adaptation in human right atrial myocytes with sinus rhythm (SR, N/n: 14/52). In mice, nNOS inhibition (N/n: 4/9) or nNOS-/- (N/n: 9/28) reduce APD50 and 90 by 53% and 35%, respectively.Ionic investigations show SMTC increases atrial IKur by 60%, Ito by 34% and IK1 by 27% with no change in ICa or IKr. Computer modelling shows mimicking SMTC-induced increases in IKur and IK1 successfully retrieves experimental phenotype. Furthermore blocking IKur abolishes the effect of nNOS inhibition on APD and APD rate-dependent adaptation (SMTC: N/n: 6/11 vs vehicle: N/n: 8/16). Additionally, In vivo atrial burst stimulation shows nNOS-/- mice exhibit 2 folds higher AF inducibility.In AF, upregulation of atrial specific miR-31 results in accelerating nNOS mRNA and protein decay. Inhibition of miR-31 recovers the SMTC suppressed APD90 (N/n: 3/12 in each group) and APD rate-dependent adaptation. These effects are NO-mediated as they are reversed by SMTC. Whereas in SR patients, increasing miR-31(N/n: 4/30) reduces nNOS, shortens APD and suppresses APD rate-dependent adaptation (N/n: 3/14).Conclusions: In human and mammalian atrial myocytes, a reduction in nNOS, mediating by upregulation of miR31, is an important factor retrieving key hallmarks of atrial electrical remodelling and contributing to the atrial phenotype begetting AF.