I. Tong Mak

Dietary magnesium intake influences circulating pro-inflammatory neuropeptide levels and loss of myocardial tolerance to postischemic stress.

Severe dietary Mg restriction (Mg9, 9% of recommended daily allowance [RDA], plasma Mg = 0.25 mM) induces a proinflammatory neurogenic response in rats (substance P [SP]), and the associated increases in oxidative stress in vivo and cardiac susceptibility to ischemla/reperfusion (I/R) injury were previously shown to be attenuated by SP receptor blockade and antioxidant treatment. The present study assessed if less severe dietary Mg restriction modulates the extent of both the neurogenic/oxidative responses in vivo and I/R injury in vitro. Male Sprague-Dawley rats maintained on Mg40 (40% RDA, plasma Mg = 0.6 mM) or Mg100 (100% RDA, plasma Mg = 0.8 mM) diets were assessed for plasma SP levels (CHEM-ELISA) during the first 3 weeks and were compared with the Mg9 group; red blood cell (RBC) glutathione and plasma malondialdehyde levels were compared at 3 weeks in Mg9, Mg20 (plasma Mg = 0.4 mM), Mg40, and Mg100 rats; and 40-min global ischemia/30-min reperfuslon hearts from 7-week-old Mg20, Mg40, and Mg100 rats were compared with respect to functional recovery (cardiac work, and diastolic, systolic, and developed pressures), tissue LDH release, and free radical production (ESR spectroscopy and α-phenyl-N-tert butyinitrone [PBN; 3 mM] spin trapping). The Mg40 diet induced smaller elevations in plasma SP (50% lower) compared with Mg9, but with a nearly identical time course. RBC glutathione and plasma malondialdehyde levels revealed a direct relationship between the severity of oxidative stress and hypomagnesemia. The dominant lipid free radical species detected in all I/R groups was the alkoxyl radical (PBN/alkoxyl: αH = 1.93 G, αN = 13.63 G); however, Mg40 and Mg20 hearts exhibited 2.7- and 3.9-fold higher alkoxyl levels, 40% and 65% greater LDH release, and lower functional recovery (Mg20 < Mg40) compared with Mg100. Our data suggest that varying dietary Mg intake directly influences the magnitude of the neurogenic/oxidative responses in vivo and the resultant myocardial tolerance to I/R Stress.

Magnesium deficiency prolongs myocardial stunning in an open-chest swine model

The effect of magnesium deficiency on postischemic myocardial dysfunction (myocardial stunning) in an open-chest swine model was studied. Twelve swine were assigned either to low magnesium diet or control diet. Myocardial stunning was assessed by measuring regional wall thickening by epicardial Doppler before and after brief occlusion (8 min) of the left anterior descending coronary artery. Serum magnesium levels decreased significantly in the experimental group only. Glutathione levels were 42.6% lower in the magnesium deficient swine than in controls. Stunning time was significantly prolonged from 32.8 +/- 3.1 min in the control group to 43.8 +/- 4.6 min in the hypomagnesemic swine. In conclusion, magnesium deficiency is associated with prolonged recovery from myocardial stunning.

Intestinal and cardiac inflammatory response shows enhanced endotoxin receptor (CD14) expression in magnesium deficiency.

Substance P is elevated in plasma and in other tissues during Mg-deficiency, and was found localised to neuronal C-fibres of cardiac and intestinal tissues, where it could promote neurogenic inflammation. Plasma prostaglandin E2 (PGE2), indicative of systemic inflammation, rose significantly (≥4 fold, p<0.01) after 1 week and remained elevated through week 2 and 3 in rat on the Mg-deficient (MgD) diet. Concomitantly, total blood glutathione decreased by 50%. Immunohistochemical staining for endotoxin (lipopolysaccaride, LPS) receptor, CD14 was prominent in macrophage-type cells in intestinal tissue; more importantly, cardiac tissue revealed both CD11b (monocyte/macrophage surface protein) and CD14 positive cells after 3 weeks in rats on MgD diet. Western blot analysis indicated a significant increase in the endotoxin receptor protein level in the 3 week MgD hearts. Since CD14 is known to be up-regulated in cells exposed to LPS, these observations suggest that prolonged Mg-deficiency results in increased intestinal permeability to bacterial products that induce the endotoxin receptor in cells localized to myocardial and intestinal tissues. These CD14 positive cells may amplify the cardiomyopathic inflammatory process by stimulating TNF-α and other pro-inflammatory cytokines. (Mol Cell Biochem 278: 53–57, 2005)

EGFR-TKI, erlotinib, causes hypomagnesemia, oxidative stress, and cardiac dysfunction: attenuation by NK-1 receptor blockade.

Abstract: To determine whether the epidermal growth factor receptor tyrosine kinase inhibitor, erlotinib may cause hypomagnesemia, inflammation, and cardiac stress, erlotinib was administered to rats (10 mg·kg−1·d−1) for 9 weeks. Plasma magnesium decreased progressively between 3 and 9 weeks (−9% to −26%). Modest increases in plasma substance P (SP) occurred at 3 (27%) and 9 (25%) weeks. Neutrophil superoxide-generating activity increased 3-fold, and plasma 8-isoprostane rose 210%, along with noticeable appearance of cardiac perivascular nitrotyrosine. The neurokinin-1 (NK-1) receptor antagonist, aprepitant (2 mg·kg−1·d−1), attenuated erlotinib-induced hypomagnesemia up to 42%, reduced circulating SP, suppressed neutrophil superoxide activity and 8-isoprostane elevations; cardiac nitrotyrosine was diminished. Echocardiography revealed mild to moderately decreased left ventricular ejection fraction (−11%) and % fractional shortening (−17%) by 7 weeks of erlotinib treatment and significant reduction (−17.5%) in mitral valve E/A ratio at week 9 indicative of systolic and early diastolic dysfunction. Mild thinning of the left ventricular posterior wall suggested early dilated cardiomyopathy. Aprepitant completely prevented the erlotinib-induced systolic and diastolic dysfunction and partially attenuated the anatomical changes. Thus, chronic erlotinib treatment does induce moderate hypomagnesemia, triggering SP-mediated oxidative/inflammation stress and mild-to-moderate cardiac dysfunction, which can largely be corrected by the administration of the SP receptor blocker.

Genomic Profiling Reveals the Potential Role of TCL1A and MDR1 Deficiency in Chemotherapy-Induced Cardiotoxicity

Background: Anthracyclines, such as doxorubicin (Adriamycin), are highly effective chemotherapeutic agents, but are well known to cause myocardial dysfunction and life-threatening congestive heart failure (CHF) in some patients. Methods: To generate new hypotheses about its etiology, genome-wide transcript analysis was performed on whole blood RNA from women that received doxorubicin-based chemotherapy and either did, or did not develop CHF, as defined by ejection fractions (EF)≤40%. Women with non-ischemic cardiomyopathy unrelated to chemotherapy were compared to breast cancer patients prior to chemo with normal EF to identify heart failure-related transcripts in women not receiving chemotherapy. Byproducts of oxidative stress in plasma were measured in a subset of patients. Results: The results indicate that patients treated with doxorubicin showed sustained elevations in oxidative byproducts in plasma. At the RNA level, women who exhibited low EFs after chemotherapy had 260 transcripts that differed >2-fold (p<0.05) compared to women who received chemo but maintained normal EFs. Most of these transcripts (201) were not altered in non-chemotherapy patients with low EFs. Pathway analysis of the differentially expressed genes indicated enrichment in apoptosis-related transcripts. Notably, women with chemo-induced low EFs had a 4.8-fold decrease in T-cell leukemia/lymphoma 1A (TCL1A) transcripts. TCL1A is expressed in both cardiac and skeletal muscle, and is a known co-activator for AKT, one of the major pro-survival factors for cardiomyocytes. Further, women who developed low EFs had a 2-fold lower level of ABCB1 transcript, encoding the multidrug resistance protein 1 (MDR1), which is an efflux pump for doxorubicin, potentially leading to higher cardiac levels of drug. In vitro studies confirmed that inhibition of MDR1 by verapamil in rat H9C2 cardiomyocytes increased their susceptibility to doxorubicin-induced toxicity. Conclusions: It is proposed that chemo-induced cardiomyopathy may be due to a reduction in TCL1A levels, thereby causing increased apoptotic sensitivity, and leading to reduced cardiac MDR1 levels, causing higher cardiac levels of doxorubicin and intracellular free radicals. If so, screening for TCL1A and MDR1 SNPs or expression level in blood, might identify women at greatest risk of chemo-induced heart failure.

Mg supplementation attenuates ritonavir-induced hyperlipidemia, oxidative stress, and cardiac dysfunction in rats

Use of protease inhibitors (PI) in HIV patients is associated with hyperlipidemia and increased risk of coronary heart disease. Chronic systemic and cardiac effects of ritonavir (RTV), a universal PI booster, and Mg supplementation were examined. RTV was administered (75 mg·kg(-1)·day(-1) po) to Lewis × Brown-Norway hybrid (LBNF1) rats for up to 8 wk; significant increases in plasma triglyceride and cholesterol occurred from 8 days to 8 wk. At 5 wk, the expression of selected hepatic genes (CYP7A1, CITED2, G6PC, and ME-1), which are key to lipid catabolism/synthesis, were altered toward lipogenesis. Dietary Mg supplementation (six-fold higher) completely reversed the altered expression of these genes and attenuated both hypertriglyceridemia and hypercholesterolemia. Neutrophils isolated from the RTV-treated rats displayed a three-fold higher basal and a twofold higher stimulated superoxide production; plasma isoprostane and red blood cell (RBC) GSSG levels were elevated two- to three-fold. All oxidative indices were normalized by Mg supplementation. After 5 wk, RTV caused significant decreases in cardiac left ventricular (LV) shortening fraction and LV ejection fraction; mitral valve early/late atrial ventricular filling (E/A) ratio was reduced accompanied by LV posterior wall thinning. Immunohistochemical staining revealed significant white blood cell (WBC) infiltration (5 wk) and prominent fibrosis (8 wk) in the RTV hearts. Mg supplementation attenuated RTV-induced declines in systolic and diastolic (improved mitral valve E/A ratio) function (>70%), lessened LV posterior wall thinning (by 75%), and substantially decreased the pathological markers. The known clinical hyperlipidemia effects of RTV can be mimicked in the LBNF1 rats; in association, systemic oxidative stress and progressive cardiac dysfunction occurred. Remarkably, Mg supplementation alone suppressed RTV-mediated hyperlipidemia, oxidative stress, and cardiac dysfunction.

d-Propranolol protects against oxidative stress and progressive cardiac dysfunction in iron overloaded rats

d-Propranolol (d-Pro: 2-8 mg·(kg body mass)(-1)·day(-1)) protected against cardiac dysfunction and oxidative stress during 3-5 weeks of iron overload (2 mg Fe-dextran·(g body mass)(-1)·week(-1)) in Sprague-Dawley rats. At 3 weeks, hearts were perfused in working mode to obtain baseline function; red blood cell glutathione, plasma 8-isoprostane, neutrophil basal superoxide production, lysosomal-derived plasma N-acetyl-β-galactosaminidase (NAGA) activity, ventricular iron content, and cardiac iron deposition were assessed. Hearts from the Fe-treated group of rats exhibited lower cardiac work (26%) and output (CO, 24%); end-diastolic pressure rose 1.8-fold. Further, glutathione levels increased 2-fold, isoprostane levels increased 2.5-fold, neutrophil superoxide increased 3-fold, NAGA increased 4-fold, ventricular Fe increased 4.9-fold; and substantial atrial and ventricular Fe-deposition occurred. d-Pro (8 mg) restored heart function to the control levels, protected against oxidative stress, and decreased cardiac Fe levels. After 5 weeks of Fe treatment, echocardiography revealed that the following were depressed: percent fractional shortening (%FS, 31% lower); left ventricular (LV) ejection fraction (LVEF, 17%), CO (25%); and aortic pressure maximum (P(max), 24%). Mitral valve E/A declined by 18%, indicating diastolic dysfunction. Cardiac CD11b+ infiltrates were elevated. Low d-Pro (2 mg) provided modest protection, whereas 4-8 mg greatly improved LVEF (54%-75%), %FS (51%-81%), CO (43%-78%), P(max) (56%-100%), and E/A >100%; 8 mg decreased cardiac inflammation. Since d-Pro is an antioxidant and reduces cardiac Fe uptake as well as inflammation, these properties may preserve cardiac function during Fe overload.

DETECTION AND MEASUREMENT OF FREE RADICAL GENERATION DURING CARDIOVASCULAR INJURY

It has been suggested thatduring myocardial ischemia short-lived free radicals such as superoxide (O 2 - ·), hydroxyl (·OH), or lipid peroxide (RO.) are produced. The evidence, however, of the participation of such radicals in cardiovascular injury has been, for the most part, based upon activation of lipid peroxidation and on myocardial protection provided by scavengers.1,2 Direct detection of such radicals, as might be possible by electron spin resonance spectroscopy (ESR) has not provided conclusive identification of radicals produced during these injuries. However, spin trapping and ESR offer an indirect technique that provides the evidence necessary to confirm the involvement of these labile intermediates. Spin trapping has been applied very successfully to studies of free radical mechanisms in a variety of chemical and photochemical reactions; however, it has only recently been applied to biological systems. The majority of the spin traps being used are molecules containing a nitroso or nitrone group which, upon addition of a labile free radical, form a stable nitroxide that is easily detected by ESR.

The EGFR tyrosine kinase inhibitor tyrphostin AG-1478 causes hypomagnesemia and cardiac dysfunction

We determined whether the epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI) N-​(3-​chlorophenyl)-​6,​7-​dimethoxy-​4-​quinazolinamine (tyrphostin AG-1478) causes hypomagnesemia and cardiac dysfunction in rats. Tyrphostin was administered (3 times per week, intraperitoneal injection, to achieve 21.4 mg·(kg body mass)(-1)·day(-1)) to normomagnesemic rats for 5 weeks. Levels of magnesium in the plasma of the tyrphostin-treated rats decreased significantly by the following amount: 17% at week 1, 27% at week 2, and 26%-35% between weeks 3 to 5. Levels of the plasma lipid peroxidation marker 8-isoprostane rose significantly: by 58% at week 1, 168% at week 3, and 113% at week 5. At week 5, blood neutrophils from the tyrphostin-treated group displayed a 2.26-fold higher basal level of O(2)(·-) generation; the ratio of oxidized glutathione (glutathione disulfide; GSSG) to reduced glutathione (GSH) in the red blood cells increased 2.5-fold. At week 5, echocardiography revealed that TKI treatment resulted in significant cardiac systolic dysfunction, with impaired diastolic function and dilated cardiomyopathy. Since hypomagnesemia alone can trigger oxidative stress and cardiac injury, we suggest that inhibition of EGFR-TK caused magnesium wasting, which partly contributed to decreased cardiac contractility.

Role of Tissue and Circulating Substance P in Cardiovascular Injury Associated with Mg-Deficiency

Hypomagnesemia is a common electrolyte deficiency found among hospitalized patients and is particularly prevalent in selected patient populations, such as alcoholics, diabetics, and those receiving diuretics and other magnesium-wasting drugs. Clinical complications as a result of magnesium deficiency were documented in a recent prospective study in which hypomagnesemia, which was present at the time of admission of critically ill patients, was associated with a statistically significant higher mortality rate [1]. Magnesium deficiency has also been associated with adverse cardiovascular conditions, such as sudden death, ventricular and atrial arrhythmias, coronary spasm, and cardiomyopathies. In one study of patients with heart disease, 45% of patients with myocardial infarction were reported to be hypomagnesemic [2].