Lerman, A.

Abstract Background Vascular aging is a key feature of cardiovascular disease. α-klotho (klotho) is a protein inversely associated with aging and may contribute to vascular repair. Lower levels of systemic klotho have been identified in humans with features of early and advanced coronary atherosclerosis. It remains unknown if circulating klotho is present within human coronary circulation and whether cardiovascular klotho retention correlates with atherosclerosis. Purpose The current study aimed to assess the presence of klotho within coronary circulation compared to systemic circulation and hypothesized that net coronary klotho retention is attenuated in patients with early age-related features of coronary artery disease (CAD). Methods A total of 42 patients that underwent clinically indicated coronary angiogram for chest pain that had no obstructive CAD underwent intravascular ultrasound (IVUS) assessment were retrospectively examined. Virtual histology assessed coronary plaque characteristics. Blood samples simultaneously collected from the proximal aorta and coronary sinus were used for klotho detection using enzyme-linked immunosorbent assay. The net gradient of klotho across cardiovascular circulation was calculated by multiplying the arteriovenous klotho gradient by basal coronary blood flow. A net positive gradient indicated less cardiovascular klotho retention; a net negative gradient indicated greater cardiovascular klotho retention. Results Overall, participants were age 54±10 years old and 76% were female. There were overall higher levels of klotho within the coronary sinus (median 0.49 ng/mL [0.15; 1.69]) compared to systemic circulation (median 0.30 ng/mL [0.16; 1.14]). Participant groups were determined by those with positive (n = 20) or negative (n = 22) cardiovascular retention of klotho. Both groups were similar in age (56 ±2 vs. 51 ±2 years old, p = 0.080), gender (72% vs. 80% female, p = 0.580), and had similar occurrences of traditional cardiovascular comorbidities. Those with less cardiovascular klotho retention in the coronary vascular bed had significantly greater degree of non-obstructive coronary artery plaque (Gensini score 3.5 vs. 0, p < 0.001) and had higher percent plaque burden (46% [35; 53] vs. 34% [28; 46], p = 0.044) than those with greater cardiovascular klotho retention. Conclusions Patients with early coronary atherosclerosis may be characterized by attenuated retention of klotho within coronary circulation. Moreover, humans appear to have higher circulating klotho in coronary circulation than in systemic circulation. The current study may support a role for mechanism or potential therapeutic target for klotho in coronary atherosclerosis in humans.

BACKGROUND Hypercholesterolemia is characterized by a coronary vasoconstrictive response to the endothelium-dependent vasodilator acetylcholine. This abnormality may be due to reduced synthesis of endothelium-derived relaxing factor and/or enhanced synthesis and release of an endothelium-derived contracting factor. Endothelin is an endothelium-derived vasoconstrictor and mitogenic peptide that is present in normal plasma, and its circulating concentrations are elevated in disease states that are characterized by abnormal endothelium-dependent relaxation to acetylcholine. The current studies were designed to test the hypotheses that experimental hypercholesterolemia results in elevation of plasma and tissue endothelin immunoreactivity and that the abnormal acetylcholine-evoked coronary vasoconstriction in the hypercholesterolemic animals is associated with further elevation of plasma endothelin. METHODS AND RESULTS Plasma concentrations and molecular forms of endothelin immunoreactivity were determined following 2% cholesterol diet for 4 months in pigs and during intracoronary acetylcholine administration. Second, we assessed the presence of endothelin in the coronary vascular wall by using immunohistochemistry. Hypercholesterolemia elevated plasma endothelin concentration and enhanced coronary artery tissue endothelin immunoreactivity. The endothelium-dependent vasodilator acetylcholine further increases plasma endothelin in hypercholesterolemia in association with coronary vasoconstriction. The predominant molecular form of endothelin in hypercholesterolemia is the biological active endothelin-1. CONCLUSIONS This study suggests a role for endothelin as an early participant and a marker for the endothelial dysfunction in hypercholesterolemia as well as a participant in the atherogenic process.

BACKGROUND Although recent investigations report the elevation of plasma endothelin (ET) in congestive heart failure (CHF), it remains unclear if this elevation is that of the biologically active peptide ET-1 or of its precursor big-ET. Furthermore, it is unclear if such elevation is associated with increased myocardial ET and if the molecular form from cardiac tissue is altered ET. Last, it remains to be established whether circulating ET is increased at the earliest stage of CHF in patients with asymptomatic left ventricular dysfunction and correlates with the magnitude of ventricular dysfunction. METHODS AND RESULTS The present study was designed to investigate concentrations and molecular forms of ET in plasma and cardiac tissue in healthy subjects and CHF patients with New York Heart Association (NYHA) class I through IV using cardiac radionuclide angiogram, cardiac myocardial biopsy, radioimmunoassay, gel permeation chromatography (GPC), and immunohistochemical staining (IHCS). Plasma ET was increased only in patients with moderate (NYHA class III) or severe (NYHA class IV) CHF compared with healthy subjects and individuals with asymptomatic (NYHA class I) or mild (NYHA class II) CHF. The elevation of circulating ET in CHF showed a negative correlation with left ventricular ejection fraction and cardiac index and a positive correlation with functional class and left ventricular end-diastolic volume index. GPC demonstrated that immunoreactive plasma ET was ET-1 in healthy subjects and both mature ET-1 and its precursor big-ET in severe CHF patients, with big-ET the predominant molecular form. Cardiac tissue concentrations and IHCS revealed ET presence in healthy atrial and ventricular tissue, which were not different in severe CHF. GPC revealed that the molecular form of cardiac ET was ET-1 in both healthy and CHF hearts. CONCLUSIONS The present study establishes for the first time that the elevation of plasma ET in severe human CHF represents principally elevation of big-ET. Second, ET is present in healthy and failing myocardia, and its activity by both immunohistochemistry and radioimmunoassay is not changed in CHF. Furthermore, the elevated plasma ET is characteristic of severe CHF and not asymptomatic or mild CHF. In addition, the degree of plasma elevation of ET correlates with the magnitude of alterations in cardiac hemodynamics and functional class. The present study confirms and extends previous investigations of ET in human CHF and establishes the evolution of circulating and local cardiac ET in the spectrum of human CHF.

BACKGROUND The endothelium possesses the ability to modulate vascular tone by the release of vasodilators and vasoconstrictors, among them endothelium-derived relaxing factor (EDRF) and endothelin (ET). Abnormalities in EDRF generation have been demonstrated in various cardiovascular pathophysiological states. Moreover, a twofold increase in plasma ET concentration was reported in these disease states. Recent in vitro studies have suggested the interaction between these two endothelium-derived substances, suggesting that imbalance between the two may contribute to alternation in vascular tone characteristic of these disease states. Thus, the hypothesis of this study was that inhibition of endogenous EDRF will enhance the vasoconstrictor response to a twofold increase in plasma ET concentrations. METHODS AND RESULTS Experiments were conducted in three groups of anesthetized dogs. In group 1, ET-1 was infused intravenously to double circulating ET concentrations. Group 2 received both ET and NG-monomethyl L-arginine (L-NMMA), a competitive inhibitor of EDRF generation, and group 3 received a continuous infusion of L-NMMA alone. Twofold increase in plasma ET concentrations was characterized by an increase in systemic and renal vasoconstriction. The inhibition of EDRF markedly enhanced the vasoconstriction to ET specifically involving the systemic, pulmonary, coronary, and renal arterial circulations. CONCLUSIONS The present study demonstrates that inhibition of endogenous EDRF augments the vasoconstrictor property of ET and supports a functional role for the balance between endothelium-derived vasodilating and vasoconstricting factors in the regulation of vascular tone.