ALDH2 polymorphism and myocardial infarction: From alcohol metabolism to redox regulation.

Acute myocardial infarction (AMI) remains a leading cause of death worldwide. Increased formation of reactive oxygen species (ROS) during the early reperfusion phase is thought to trigger lipid peroxidation and disrupt redox homeostasis, leading to myocardial injury. Whilst the mitochondrial enzyme aldehyde dehydrogenase 2 (ALDH2) is chiefly recognised for its central role in ethanol metabolism, substantial experimental evidence suggests an additional cardioprotective role for ALDH2 independent of alcohol intake, which mitigates myocardial injury by detoxifying breakdown products of lipid peroxidation including the reactive aldehydes, malondialdehyde (MDA) and 4-hydroxynonenal (4-HNE). Epidemiological evidence suggests that an ALDH2 mutant variant with reduced activity that is highly prevalent in the East Asian population increases AMI risk. Additional studies have uncovered a strong association between coronary heart disease and this ALDH2 mutant variant. It appears this enzyme polymorphism (in particular, in ALDH2*2/2 carriers) has the potential to have wide-ranging effects on thiol reactivity, redox tone and therefore numerous redox-related signaling processes, resilience of the heart to cope with lifestyle-related and environmental stressors, and the ability of the whole body to achieve redox balance. In this review, we summarize the journey of ALDH2 from a mitochondrial reductase linked to alcohol metabolism, via pre-clinical studies aimed at stimulating ALDH2 activity to reduce myocardial injury to clinical evidence for its protective role in the heart.

Cysteine hydropersulfide reduces lipid peroxidation and protects against myocardial ischaemia-reperfusion injury - Are endogenous persulfides mediators of ischaemic preconditioning?

Earlier studies revealed the presence of cysteine persulfide (CysSSH) and related polysulfide species in various mammalian tissues. CysSSH has both antioxidant and oxidant properties, modulates redox-dependent signal transduction and has been shown to mitigate oxidative stress. However, its functional relevance in the setting of myocardial ischaemia-reperfusion injury (IRI) remains unknown. The present study was undertaken to (1) study the dynamics of production and consumption of persulfides under normoxic and hypoxic conditions in the heart, and (2) determine whether exogenous administration of the CysSSH donor, cysteine trisulfide (Cys-SSS-Cys) at the onset of reperfusion rescues functional impairment and myocardial damage by interfering with lipid peroxidation. Utilising a well-established ex vivo Langendorff murine model, we here demonstrate that endogenous tissue concentrations of CysSSH are upregulated when oxygen supply is compromised (global myocardial ischaemia) and rapidly restored to baseline levels upon reperfusion, suggestive of active regulation. In a separate set of experiments, exogenous administration of Cys-SSS-Cys for 10 min at the onset of reperfusion was found to decrease malondialdehyde (MDA) concentrations, formation of 4-hydroxynonenal (4-HNE) protein adducts and rescue the heart from injury. Cys-SSS-Cys also restored post-ischaemic cardiac function, improving both coronary flow and left ventricular developed pressure (LVDP). Taken together, these results support the notion that endogenous CysSSH plays an important role as a "redox preconditioning" agent to combat the oxidative insult in myocardial IRI.

The Use of Wire Myography to Investigate Vascular Tone and Function.

Wire myography enables the investigation of vascular tone and function of small vessels. The vessel of interest is harvested from the experimental model of choice, and then mounted as ring preparations onto a four-channel wire myograph. This technique enables ex vivo measurements of isometric response of vessels to different pharmacological agents. Here we describe in detail how to dissect, mount, and normalize vessels for the wire myography technique. We will also provide examples of how to construct concentration-response curves to a contractile and vasodilatory pharmacological agent.

Nitrite and myocardial ischaemia reperfusion injury. Where are we now?

Cardiovascular disease remains the leading cause of death worldwide despite major advances in technology and treatment, with coronary heart disease (CHD) being a key contributor. Following an acute myocardial infarction (AMI), it is imperative that blood flow is rapidly restored to the ischaemic myocardium. However, this restoration is associated with an increased risk of additional complications and further cardiomyocyte death, termed myocardial ischaemia reperfusion injury (IRI). Endogenously produced nitric oxide (NO) plays an important role in protecting the myocardium from IRI. It is well established that NO mediates many of its downstream functions through the 'canonical' NO-sGC-cGMP pathway, which is vital for cardiovascular homeostasis; however, this pathway can become impaired in the face of inadequate delivery of necessary substrates, in particular L-arginine, oxygen and reducing equivalents. Recently, it has been shown that during conditions of ischaemia an alternative pathway for NO generation exists, which has become known as the 'nitrate-nitrite-NO pathway'. This pathway has been reported to improve endothelial dysfunction, protect against myocardial IRI and attenuate infarct size in various experimental models. Furthermore, emerging evidence suggests that nitrite itself provides multi-faceted protection, in an NO-independent fashion, against a myriad of pathophysiologies attributed to IRI. In this review, we explore the existing pre-clinical and clinical evidence for the role of nitrate and nitrite in cardioprotection and discuss the lessons learnt from the clinical trials for nitrite as a perconditioning agent. We also discuss the potential future for nitrite as a pre-conditioning intervention in man.

Association of reduced inner retinal thicknesses with chronic kidney disease.

BACKGROUND: Tissue derived biomarkers may offer utility as indicators of accumulated damage. Reduced thickness of retinal neuronal tissue and the vascular choroid have previously been associated with vascular damage and diabetes. We evaluated associations between retinal thickness, retinal microvascular and choroidal measures, and renal function in a population with a high burden of comorbidity. METHODS: Participants were recruited from nuclear cardiology or renal medicine clinics. Retinal and choroidal thickness were measured from spectral-domain optical coherence tomograms. Retinal microvascular parameters were assessed from digital fundus photographs using a semi-automated software package. MAIN OUTCOME MEASURE: Chronic kidney disease (CKD) categorised as: CKD stages 1-2, eGFR ≥60 ml/min/1.73m2; CKD stage 3, eGFR 30-59 ml/min/1.73m2, and CKD stages 4-5, eGFR ≤29 ml/min/1.73m2. RESULTS: Participants (n = 241) had a mean age of 65 years and a mean eGFR of 66.9 ml/min/1.73m2. Thirty-nine % of the cohort had diabetes and 27% were using diuretics. Thinning of the inner retina and changes to its microvascular blood supply were associated with lower eGFR and CKD stages 4 and 5, while no associations were found between the outer retinal layers or their choroidal blood supply and CKD of any stage. These associations remained following adjustment for age, mean arterial blood pressure, diabetes status, low-density lipoprotein, body mass index, and sex. CONCLUSIONS: Inner retinal thinning and retinal microvascular variation is associated with advanced CKD (stages 4 & 5) independent of important confounding factors, but not with earlier stage CKD (stage 3) and, therefore, its utility as a biomarker for early CKD is not supported in this study.

Long-lasting blood pressure lowering effects of nitrite are NO-independent and mediated by hydrogen peroxide, persulfides, and oxidation of protein kinase G1α redox signalling.

AIMS: Under hypoxic conditions, nitrite (NO2-) can be reduced to nitric oxide (NO) eliciting vasorelaxation. However, nitrite also exerts vasorelaxant effects of potential therapeutic relevance under normal physiological conditions via undetermined mechanisms. We, therefore, sought to investigate the mechanism(s) by which nitrite regulates the vascular system in normoxia and, specifically, whether the biological effects are a result of NO generation (as in hypoxia) or mediated via alternative mechanisms involving classical downstream targets of NO [e.g. effects on protein kinase G1α (PKG1α)]. METHODS AND RESULTS: Ex vivo myography revealed that, unlike in thoracic aorta (conduit vessels), the vasorelaxant effects of nitrite in mesenteric resistance vessels from wild-type (WT) mice were NO-independent. Oxidants such as H2O2 promote disulfide formation of PKG1α, resulting in NO- cyclic guanosine monophosphate (cGMP) independent kinase activation. To explore whether the microvascular effects of nitrite were associated with PKG1α oxidation, we used a Cys42Ser PKG1α knock-in (C42S PKG1α KI; 'redox-dead') mouse that cannot transduce oxidant signals. Resistance vessels from these C42S PKG1α KI mice were markedly less responsive to nitrite-induced vasodilation. Intraperitoneal (i.p.) bolus application of nitrite in conscious WT mice induced a rapid yet transient increase in plasma nitrite and cGMP concentrations followed by prolonged hypotensive effects, as assessed using in vivo telemetry. In the C42S PKG1α KI mice, the blood pressure lowering effects of nitrite were lower compared to WT. Increased H2O2 concentrations were detected in WT resistance vessel tissue challenged with nitrite. Consistent with this, increased cysteine and glutathione persulfide levels were detected in these vessels by mass spectrometry, matching the temporal profile of nitrite's effects on H2O2 and blood pressure. CONCLUSION: Under physiological conditions, nitrite induces a delayed and long-lasting blood pressure lowering effect, which is NO-independent and occurs via a new redox mechanism involving H2O2, persulfides, and PKG1α oxidation/activation. Targeting this novel pathway may provide new prospects for anti-hypertensive therapy.

Serum amyloid A levels are associated with polymorphic variants in the serum amyloid A 1 and 2 genes.

BACKGROUND: Serum amyloid A (SAA) is secreted by liver hepatocytes in response to increased inflammation whereupon it associates with high-density lipoprotein (HDL) and alters the protein and lipid composition of HDL negating some of its anti-atherogenic properties. AIMS: To identify variants within the SAA gene that may be associated with SAA levels and/or cardiovascular disease (CVD). METHODS: We identified exonic variants within the SAA genes by deoxyribonucleic acid (DNA) Sanger sequencing. We tested the association between SAA variants and serum SAA levels in 246 individuals with and without CVD. RESULTS: Increased SAA was associated with rs2468844 (beta [ß] = 1.73; confidence intervals [CI], 1.14-1.75; p = 0.01), rs1136747 (ß = 1.53 (CI, 1.11-1.73); p = 0.01) and rs149926073 (ß = 3.37 (CI, 1.70-4.00); p = 0.02), while rs1136745 was significantly associated with decreased SAA levels (ß = 0.70 (CI, 0.53-0.94); p = 0.02). Homozygous individuals with the SAA1.3 haplotype had significantly lower levels of SAA compared with those with SAA1.1 or SAA1.5 (ß = 0.43 (CI, 0.22-0.85); p = 0.02) while SAA1.3/1.5 heterozygotes had significantly higher SAA levels compared with those homozygous for SAA1.1 (ß = 2.58 (CI, 1.19-5.57); p = 0.02). CONCLUSIONS: We have identified novel genetic variants in the SAA genes associated with SAA levels, a biomarker of inflammation and chronic disease. The utility of SAA as a biomarker for inflammation and chronic disease may be influenced by underlying genetic variation in baseline levels.

Type 2 Diabetes in Young Females Results in Increased Serum Amyloid A and Changes to Features of High Density Lipoproteins in Both HDL2 and HDL3.

Persons with type 2 diabetes mellitus (T2DM) have an elevated risk of atherosclerosis. High-density lipoproteins (HDL) normally protect against cardiovascular disease (CVD), but this may be attenuated by serum amyloid A (SAA). In a case-control study of young females, blood samples were compared between subjects with T2DM (n = 42) and individuals without T2DM (n = 42). SAA and apolipoprotein AI (apoAI) concentrations, paraoxonase-1 (PON-1), cholesteryl ester transfer protein (CETP), and lecithin-cholesterol acyltransferase (LCAT) activities were measured in the serum and/or HDL2 and HDL3 subfractions. SAA concentrations were higher in T2DM compared to controls: serum (30 mg/L (17, 68) versus 15 mg/L (7, 36); p = 0.002), HDL2 (1.0 mg/L (0.6, 2.2) versus 0.4 mg/L (0.2, 0.7); p < 0.001), and HDL3, (13 mg/L (8, 29) versus 6 mg/L (3, 13); p < 0.001). Serum-PON-1 activity was lower in T2DM compared to that in controls (38,245 U/L (7025) versus 41,109 U/L (5690); p = 0.043). CETP activity was higher in T2DM versus controls in HDL2 (232.6 µmol/L (14.1) versus 217.1 µmol/L (25.1); p = 0.001) and HDL3 (279.5 µmol/L (17.7) versus 245.2 µmol/L (41.2); p < 0.001). These results suggest that individuals with T2DM have increased SAA-related inflammation and dysfunctional HDL features. SAA may prove to be a useful biomarker in T2DM given its association with elevated CVD risk.