Copper excess, zinc deficiency, and cognition loss in Alzheimer's disease

In this special issue about biofactors causing cognitive impairment, we present evidence for and discuss two such biofactors. One is excess copper, causing neuronal toxicity. The other is zinc deficiency, causing neuronal damage. We present evidence that Alzheimer's disease (AD) has become an epidemic in developed, but not undeveloped, countries and that the epidemic is a new disease phenomenon, beginning in the early 1900s and exploding in the last 50 years. This leads to the conclusion that something in the developed environment is a major risk factor for AD. We hypothesize that the factor is inorganic copper, leached from the copper plumbing, the use of which coincides with the AD epidemic. We present a web of evidence supporting this hypothesis. Regarding zinc, we have shown that patients with AD are zinc deficient when compared with age-matched controls. Zinc has critical functions in the brain, and lack of zinc can cause neuronal death. A nonblinded study about 20 years ago showed considerable improvement in AD with zinc therapy, and a mouse AD model study also showed significant cognitive benefit from zinc supplementation. In a small blinded study we carried out, post hoc analysis revealed that 6 months of zinc therapy resulted in significant benefit relative to placebo controls in two cognitive measuring systems. These two factors may be linked in that zinc therapy significantly reduced free copper levels. Thus, zinc may act by lowering copper toxicity or by direct benefit on neuronal health, or both.     

Protein glycation, oxidation and nitration adduct residues and free adducts of cerebrospinal fluid in Alzheimer's disease and link to cognitive impairment

Increased damage to proteins by glycation, oxidation and nitration has been implicated in neuronal cell death leading to Alzheimer's disease (AD). Protein glycation, oxidation and nitration adducts are consequently formed. Quantitative screening of these adducts in CSF may provide a biochemical indicator for the diagnosis of AD. To assess this, we measured 11 glycation adducts, three oxidation adducts and a nitration adduct, determining both protein adduct residues and free adducts, in CSF samples of age-matched normal healthy subjects (n = 18) and subjects with Alzheimer's disease (n = 32). In CSF protein, the concentrations of 3-nitrotyrosine, N(epsilon)-carboxymethyl-lysine, 3-deoxyglucosone-derived hydroimidazolone and N-formylkynurenine residues were increased in subjects with Alzheimer's disease. In CSF ultrafiltrate, the concentrations of 3-nitrotyrosine, methylglyoxal-derived hydroimidazolone and glyoxal-derived hydroimidazolone free adducts were also increased. The Mini-Mental State Examination (MMSE) score correlated negatively with 3-nitrotyrosine residue concentration (p < 0.05), and the negative correlation with fructosyl-lysine residues just failed to reach significance (p = 0.052). Multiple linear regression gave a regression model of the MMSE score on 3-nitrotyrosine, fructosyl-lysine and N(epsilon)-carboxyethyl-lysine residues with p-values of 0.021, 0.031 and 0.052, respectively. These findings indicate that protein glycation, oxidation and nitration adduct residues and free adducts were increased in the CSF of subjects with Alzheimer's disease. A combination of nitration and glycation adduct estimates of CSF may provide an indicator for the diagnosis of Alzheimer's disease.     

Quantitative measurement of specific biomarkers for protein oxidation, nitration and glycation in Arabidopsis leaves

Higher plants are continually exposed to reactive oxygen and nitrogen species during their lives. Together with glucose and reactive dicarbonyls, these can modify proteins spontaneously, leading to protein oxidation, nitration and glycation. These reactions have the potential to damage proteins and have an impact on physiological processes. The levels of protein oxidation, nitration and glycation adducts were assayed, using liquid chromatography coupled with tandem mass spectrometry, in total leaf extracts over a diurnal cycle and when exposed to conditions that promote oxidative stress. Changes in the levels of oxidation, glycation and nitration adducts were found between the light and dark phases under non-stress conditions. A comparison between wild-type plants and a mutant lacking peptide methionine sulfoxide reductase ( pmsr2-1 ) showed increased protein oxidation, nitration and glycation of specific amino acid residues during darkness in pmsr2-1 . Short-term excess light exposure, which promoted oxidative stress, led to increased protein glycation, specifically by glyoxal. This suggested that any increased oxidative damage to proteins was within the repair capacity of the plant. The methods developed here provide the means to simultaneously detect a range of protein oxidation, nitration and glycation adducts within a single sample. Thus, these methods identify a range of biomarkers to monitor a number of distinct biochemical processes that have an impact on the proteome and therefore the physiological state of the plant.     

Detection of sequence-specific tyrosine nitration of manganese SOD and SERCA in cardiovascular disease and aging

Nitration of protein tyrosine residues (nY) is a marker of oxidative stress and may alter the biological activity of the modified proteins. The aim of this study was to develop antibodies toward site-specific nY-modified proteins and to use histochemistry and immunoblotting to demonstrate protein nitration in tissues. Affinity-purified polyclonal antibodies toward peptides with known nY sites in MnSOD nY-34 and of two adjacent nY in the sarcoplasmic endoplasmic reticulum calcium ATPase (SERCA2 di-nY-294,295) were developed. Kidneys from rats infused with ANG II with known MnSOD nY and aorta from atherosclerotic rabbits and aging rat skeletal and cardiac sarcoplasmic reticulum with known SERCA di-nY were used for positive controls. Staining for MnSOD nY-34 was most intense in distal renal tubules and collecting ducts. Staining of atherosclerotic aorta for SERCA2 di-nY was most intense in atherosclerotic plaques. Aging rat skeletal muscle and atherosclerotic aorta and cardiac atrium from human diabetic patients also stained positively. Staining was decreased by sodium dithionite, which chemically reduces nitrotyrosine to aminotyrosine, and the antigenic nY-peptide blocked staining for each respective nY site but not for the other. As previously demonstrated, immunoblotting failed to detect these modified proteins in whole tissue lysates but did when the proteins were concentrated. Immunohistochemical staining for specific nY-modified tyrosine residues offers the ability to assess the effects of oxidant stress associated with pathological conditions on individual proteins whose function may be affected in specific tissue sites.     

The role of advanced glycation end products in retinal ageing and disease

The retina is exposed to a lifetime of potentially damaging environmental and physiological factors that make the component cells exquisitely sensitive to age-related processes. Retinal ageing is complex and a raft of abnormalities can accumulate in all layers of the retina. Some of this pathology serves as a sinister preamble to serious conditions such as age-related macular degeneration (AMD) which remains the leading cause of irreversible blindness in the Western world.     

Niacin and cholesterol: role in cardiovascular disease (review)

Niacin has been widely used as a pharmacologic agent to regulate abnormalities in plasma lipid and lipoprotein metabolism and in the treatment of atherosclerotic cardiovascular disease. Although the use of niacin in the treatment of dyslipidemia has been reported as early as 1955, only recent studies have yielded an understanding about the cellular and molecular mechanism of action of niacin on lipid and lipoprotein metabolism. In brief, the beneficial effect of niacin to reduce triglycerides and apolipoprotein-B containing lipoproteins (e.g., VLDL and LDL) are mainly through: a) decreasing fatty acid mobilization from adipose tissue triglyceride stores, and b) inhibiting hepatocyte diacylglycerol acyltransferase and triglyceride synthesis leading to increased intracellular apo B degradation and subsequent decreased secretion of VLDL and LDL particles. The mechanism of action of niacin to raise HDL is by decreasing the fractional catabolic rate of HDL-apo AI without affecting the synthetic rates. Additionally, niacin selectively increases the plasma levels of Lp-AI (HDL subfraction without apo AII), a cardioprotective subfraction of HDL in patients with low HDL. Using human hepatocytes (Hep G2 cells) as an in vitro model system, recent studies indicate that niacin selectively inhibits the uptake/removal of HDL-apo AI (but not HDL-cholesterol ester) by hepatocytes, thereby increasing the capacity of retained HDL-apo AI to augment cholesterol efflux through reverse cholesterol transport pathway. The studies discussed in this review provide evidence to extend the role of niacin as a lipid-lowering drug beyond its role as a vitamin.     

Emerging role of SIRT3 in endothelial metabolism,angiogenesis, and cardiovascular disease

Sirtuin 3 (SIRT3) a mitochondrial enzyme that plays an important role in energy homeostasis, cardiac remodeling, and heart failure (HF). The expression of SIRT3 declines with advanced age, cardiovascular, and metabolic diseases. Accumulating evidence suggests that SIRT3 plays a critical role in protecting the heart from cardiac hypertrophy, cardiac dysfunction associated with HF, and in the protection of cardiac cells from stress-mediated cell death. Clinical studies have demonstrated that HF with preserved ejection fraction (HFpEF) in patients present with abnormalities in coronary microcirculation related to endothelial dysfunction and coronary microvascular rarefaction. Although SIRT3-mediated regulation of mitochondrial homeostasis and heart function has been intensively investigated, the effect of SIRT3 on endothelial cell (EC) glycolytic metabolism and microvascular function has not been well studied. ECs utilize glycolysis for generating ATP rather than oxidative phosphorylation to maintain their normal functions and promote angiogenesis and EC–cardiomyocyte interactions. Emerging evidence indicates that SIRT3 is involved in the regulation of endothelial metabolism and angiogenesis and thus affects the development of cardiovascular diseases associated with aging. This review will discuss the current knowledge of SIRT3 and its functional role on endothelial metabolism, cardiac function, and cardiovascular diseases.     

Endothelin and endothelin antagonists: Potential role in cardiovascular and renal disease

Endothelin-1 is a recently discovered peptide mainly released from endothelial cells. Hypoxia and ischemia as well as numerous factors such as angiotensin 11, thrombin and transforming growth factor ₁ stimulate the fomation of the peptide. On the other hand the synthesis of endothelin is inhibited by nitric oxide and atrial natriuretic peptide via the formation of cyclic guanosine monophosphate. Released from endothelial cells endothelin-1 mediates transient vasodilation followed by a profound and longlasting vasoconstriction. Endothelin is also a mitogen for smooth muscle proliferation. Endothelins exert their biological effects via activation of specific receptors. Two different receptors have been cloned from mammalian tissues (ET_A and ET_B receptors). On vascular smooth muscle cells both receptors mediate contractions. Endothelial cells only express ET_B receptors linked to the formation of nitric oxide and/or prostacyclin formation. Increased plasma concentrations of endothelin-1 have been described in a variety of diseases such as pulmonary hypertension, arteriosclerosis, renal failure, acute coronary syndromes, heart failure, migraine and vascular diseases.Recently an increasing number of endothelin receptor antagonists have been synthetized, which have been shown to inhibit endothelin-mediated vasoconstriction. Clinical studies are now ongoing to elucidate the pathophysiologic role of endothelin and the potential benefit of the blockade of the system in different disease states.     

Vitamin A and lipid peroxidation: effect of retinol deficiency

The effect of alimentary vitamin A deficiency on some parameters of lipid peroxidation (LPO) in young rats was studied. It was found that under vitamin A deficiency the content of diene conjugates in liver homogenates and microsomes diminishes, whereas that of malonic aldehyde in small intestinal mucosa, liver and testis homogenates is unaffected. However, the malonic aldehyde production in liver homogenates and microsomes decreases after 60 min incubation at 37 degrees C without addition of prooxidants. At the same time, enzymatic NADPH-dependent and nonenzymatic ascorbate-dependent LPO in liver microsomes of vitamin A-deficient rats does not change significantly. The decrease of LPO intensity in vitamin A-deficient animals may be due to the reduced content in liver microsomes of the main LPO substrates, i.e., arachidonic and linoleic acids, as well as to the decrease of cytochrome P-450 level in rat liver.     

Protein and lipid nitration: role in redox signaling and injury

Protein and lipid nitration represent novel footprints of oxidative and nitrative stress processes. In this review, we first discuss the mechanisms of formation of protein 3-nitrotyrosine and nitrated fatty acids as well as their key biological and signaling actions. Elevation of protein 3-nitrotyrosine levels is associated to tissue injury, and some specific nitrated proteins play a causative role in disease progression; on the other hand, the substantiation on the role of tyrosine nitration on redox signaling is rather scarce. Herein, we also provide evidence to support that the nitration of lipids (i.e. to nitrofatty acids) results in the formation of novel endogenous modulators of redox processes, partially counteracting pro-inflammatory effects of oxidant exposure.     

The role of intestinal microbiota in cardiovascular disease

Accumulating evidence has indicated that intestinal microbiota is involved in the development of various human diseases, including cardiovascular diseases (CVDs). In the recent years, both human and animal experiments have revealed that alterations in the composition and function of intestinal flora, recognized as gut microflora dysbiosis, can accelerate the progression of CVDs. Moreover, intestinal flora metabolizes the diet ingested by the host into a series of metabolites, including trimethylamine N‐oxide, short chain fatty acids, secondary bile acid and indoxyl sulfate, which affects the host physiological processes by activation of numerous signalling pathways. The aim of this review was to summarize the role of gut microbiota in the pathogenesis of CVDs, including coronary artery disease, hypertension and heart failure, which may provide valuable insights into potential therapeutic strategies for CVD that involve interfering with the composition, function and metabolites of the intestinal flora.     

Autophagy following heat stress: the role of aging and protein nitration

Stress can originate from a variety of sources (e.g., physical, chemical, etc.,) and cause protein denaturation, DNA damage and possibly death. In an effort to prevent such deleterious consequences, most organisms possess one or more ways to counteract or even prevent the harmful effect(s) from a given stressor. Such compensation by an organism is known as a stress response; this involves inhibition of housekeeping genes and subsequent activation of genes associated with the stress response. One of the most widely studied groups of stress response genes is a family of molecular chaperones known as heat shock proteins (HSPs). Work from our laboratory agrees with many other studies showing an age-related decline in stress-induced synthesis of HSPs. A decline in the availability and/or function of HSPs with age can lead to accumulation of damaged proteins, which in turn damages cells. Recently, our laboratory found a significant increase in mitochondrial damage as well as evidence of increased autophagy in rat hepatocytes following heat stress. These results, along with findings of increased protein nitration with age, suggest a major role for reactive nitrogen species (RNS) in both the decline in HSP induction and increased hepatocyte pathology observed in old rats following heat stress.     

Mineralocorticoid receptors in immune cells: Emerging role in cardiovascular disease

Mineralocorticoid receptors (MRs) contribute to the pathophysiology of hypertension and cardiovascular disease in humans. As such, MR antagonists improve cardiovascular outcomes but the molecular mechanisms remain unclear. The actions of the MR in the kidney to increase blood pressure are well known, but the recent identification of MRs in immune cells has led to novel discoveries in the pathogenesis of cardiovascular disease that are reviewed here. MR regulates macrophage activation to the pro-inflammatory M1 phenotype and this process contributes to the pathogenesis of cardiovascular fibrosis in response to hypertension and to outcomes in mouse models of stroke. T lymphocytes have recently been implicated in the development of hypertension and cardiovascular fibrosis in mouse models. MR activation in vivo promotes T lymphocyte differentiation to the pro-inflammatory Th1 and Th17 subsets while decreasing the number of anti-inflammatory T regulatory lymphocytes. The mechanism likely involves activation of MR in antigen presenting dendritic cells that subsequently regulate Th1/Th17 polarization by production of cytokines. Alteration of the balance between T helper and T regulatory lymphocytes contributes to the pathogenesis of hypertension and atherosclerosis and the associated complications. B lymphocytes also express the MR and specific B lymphocyte-derived antibodies modulate the progression of atherosclerosis. However, the role of MR in B lymphocyte function remains to be explored. Overall, recent studies of MR in immune cells have identified new mechanisms by which MR activation may contribute to the pathogenesis of organ damage in patients with cardiovascular risk factors. Conversely, inhibition of leukocyte MR may contribute to the protective effects of MR antagonist drugs in cardiovascular patients. Further understanding of the role of MR in leukocyte function could yield novel drug targets for cardiovascular disease.     

The role of apolipoprotein E in neurodegeneration and cardiovascular disease

Apolipoprotein E (ApoE) is an abundant plasma protein that interacts with low density lipoprotein receptors and other proteins, participating in the transport of cholesterol and lipids. Research has revealed many other roles for this multifunctional protein. ApoE is polymorphic and exists in three major isoforms: ApoE2, ApoE3 (the most common isoform) and ApoE4, which differ by only one amino acid, at positions 112 and 158. The altered binding to lipids and receptors by ApoE isoforms E2 and E4 results in an elevated risk for neurological, cerebrovascular and cardiovascular pathologies. Most notably, ApoE4 is associated with an elevated risk (relative to E3) for Alzheimer’s disease. The application of mass spectrometry for genotyping and also direct measurement of ApoE protein isoforms is a recent development and is well suited to high-throughput applications. The precise quantification of protein isoforms will allow better characterization of effects resulting from heterozygous APOE genotypes.     

The role of antimicrobial peptides in cardiovascular physiology and disease

Antimicrobial peptides are natural peptide antibiotics, existing ubiquitously in both plant and animal kingdoms. They exhibit broad-spectrum antimicrobial activity and play an important role in host defense against invading microbes. Recently, these peptides have been shown to possess activities unrelated to direct microbial killing and be involved in the complex network of immune responses and inflammation. Thus, their role has now broadened beyond that of endogenous antibiotics. Because of their wide involvement in inflammatory response and the emerging role of inflammation in atherosclerosis, antimicrobial peptides have been proposed to represent an important link between inflammation and the pathogenesis of atherosclerotic cardiovascular diseases. This review highlights recent findings that support a role of these peptides in cardiovascular physiology and disease.