Abstract-Apelin receptors, present on vascular smooth muscle cells, endothelium, and cardiomyocytes, are activated by the family of apelin peptides to elicit cardiovascular effects in experimental animals, but functional activity in humans has not been studied in detail. We detected low levels of apelin immunoreactivity in plasma of volunteers consistent with an autocrine/paracrine action and detected apelin immunoreactivity in the supernatant from human cultured endothelial cells. We found that [Pyr 1 ]apelin-13 was the predominant isoform in cardiac tissue from patients with coronary artery disease. We tested the hypothesis that apelins have vascular and cardiac actions in human tissues in vitro and compared responses to [Pyr 1 ]apelin-13, apelin-13, and apelin-36. In endothelium-intact mammary artery, all 3 of the apelins induced concentration-dependent vasodilatation with comparable potency (EC 50 : 0.6 to 1.6 nM; maximum response: 40% to 50%). Vasodilatation was abolished after endothelial removal or preincubation with indomethacin but was unaffected by preincubation with N G -nitro-L-arginine methyl ester, indicating involvement of prostanoids but not NO in dilatation by apelins in this patient group. Apelins were potent constrictors of endothelium-denuded saphenous vein (EC 50 : 0.6 to 1.6 nM; maximum response: 17% to 26%) and mammary artery ([Pyr 1 ]apelin-13; EC 50 : 0.2 nM; maximum response: 29%). In paced atrial strips, all 3 of the peptides increased the force of contraction with subnanomolar potencies (EC 50 : 40 to 125 pM). For the first time, we demonstrate that the 3 principal forms of apelin have comparable potency and efficacy in human cardiovascular tissues. Apelins are potent endothelium-dependent vasodilators acting via a prostanoid-dependent mechanism; however, removal of the endothelium revealed direct vasoconstrictor actions in both the artery and vein. Furthermore, in human cardiac tissue, the apelin peptides are among the most potent endogenous positive inotropic agents yet reported. A pelin peptides, derived from a single gene, activate the G protein-coupled receptor APJ that has high sequence similarity with the angiotensin II type 1 receptor but does not bind angiotensin II. 1,2 The precursor, preproapelin, 2-4 contains several proteolytic cleavage sites, 3 predicting formation of C-terminal peptides, including apelin-36, apelin-13, and [Pyr 1 ]apelin-13, 2,3,5 but the endogenous isoforms in humans remain to be identified. APJ modulates fluid homeostasis 4 -6 and acts as coreceptor for HIV infection, 7,8 and apelin is a novel adipokine 9 -11 with an emerging role in the cardiovascular system. 6,12-38 A role for apelin in human cardiovascular disease has been proposed; importantly, mRNA microarray analysis of Ͼ12 000 genes identified APJ as the most consistently and significantly increased gene in heart failure patients after mechanical offloading. 12 Circulating levels of apelin may be unchanged 13 or raised 12 in early stages of heart failure but were normalized 12 or decreased...

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Background and purpose: The aim of this study was to determine whether the apelin/APJ system is altered in human cardiovascular disease by investigating whether the expression of apelin or its receptor is altered at the protein level. Experimental approach: Radioligand binding studies were used to determine apelin receptor density in human cardiac tissues. Apelin peptide levels in cardiovascular tissues were determined by radioimmunoassay. In vitro pharmacology was used to assess vasoactive properties of apelin in human coronary artery. Localization of apelin and its receptor in coronary artery was determined using immunohistochemistry. Key results: Apelin receptor density was significantly decreased in left ventricle from patients with dilated cardiomyopathy or ischaemic heart disease compared with controls, but apelin peptide levels remained unchanged. Apelin was up-regulated in human atherosclerotic coronary artery and this additional peptide localized to the plaque, colocalizing with markers for macrophages and smooth muscle cells. Apelin potently constricted human coronary artery. Conclusions and implications:We have detected changes in the apelin/APJ system in human diseased cardiac and vascular tissue. The decrease in receptor density in heart failure may limit the positive inotropic actions of apelin, contributing to contractile dysfunction. The contribution of the increased apelin levels in atherosclerotic coronary artery to disease progression remains to be determined. These data suggest a potential role for the apelin/APJ system in human cardiovascular disease.

The apelin receptor (APJ) is a class A G-protein-coupled receptor (GPCR) and is a putative target for the treatment of cardiovascular and metabolic diseases. Apelin-13 (NH₂-QRPRLSHKGPMPF-COOH) is a vasoactive peptide and one of the most potent endogenous inotropic agents identified to date. We report the design and discovery of a novel APJ antagonist. By using a bivalent ligand approach, we have designed compounds with two 'affinity' motifs and a short series of linker groups with different conformational and non-bonded interaction properties. One of these, cyclo(1-6)CRPRLC-KH-cyclo(9-14)CRPRLC is a competitive antagonist at APJ. Radioligand binding in CHO cells transfected with human APJ gave a K(i) value of 82 nM, competition binding in human left ventricle gave a K(D) value of 3.2 μM, and cAMP accumulation assays in CHO-K1-APJ cells gave a K(D) value of 1.32 μM.

There is a wide variation in laboratory practice with regard to implementation and review of internal quality control (IQC). A poor approach can lead to a spectrum of scenarios from validation of incorrect patient results to over investigation of falsely rejected analytical runs. This article will provide a practical approach for the routine clinical biochemistry laboratory to introduce an efficient quality control system that will optimise error detection and reduce the rate of false rejection. Each stage of the IQC system is considered, from selection of IQC material to selection of IQC rules, and finally the appropriate action to follow when a rejection signal has been obtained. The main objective of IQC is to ensure day-to-day consistency of an analytical process and thus help to determine whether patient results are reliable enough to be released. The required quality and assay performance varies between analytes as does the definition of a clinically significant error. Unfortunately many laboratories currently decide what is clinically significant at the troubleshooting stage. Assay-specific IQC systems will reduce the number of inappropriate sample-run rejections compared with the blanket use of one IQC rule. In practice, only three or four different IQC rules are required for the whole of the routine biochemistry repertoire as assays are assigned into groups based on performance. The tools to categorise performance and assign IQC rules based on that performance are presented. Although significant investment of time and education is required prior to implementation, laboratories have shown that such systems achieve considerable reductions in cost and labour.

Context In primary aldosteronism, cosecretion of cortisol may alter cortisol-derived adrenal venous sampling indices. Objective To identify whether cortisol cosecretion in primary aldosteronism alters adrenal venous sampling parameters and interpretation. Design Retrospective case–control study Setting A tertiary referral center Patients 144 adult patients with primary aldosteronism who had undergone both adrenocorticotropic hormone-stimulated adrenal venous sampling and dexamethasone suppression testing between 2004 and 2018. Main Outcome Measures Adrenal venous sampling indices including adrenal vein aldosterone/cortisol ratios and the selectivity, lateralization, and contralateral suppression indices. Results 21 (14.6%) patients had evidence of cortisol cosecretion (defined as a failure to suppress cortisol to ≤50 nmol/L post dexamethasone). Patients with evidence of cortisol cosecretion had a higher inferior vena cava cortisol concentration (P = .01) than those without. No difference was observed between the groups in terms of selectivity index, lateralization index, lateralization of aldosterone excess, or adrenal vein cannulation rate. Conclusions Cortisol cosecretion alters some parameters in adrenocorticotrophic hormone-stimulated adrenal venous sampling but does not result in alterations in patient management.

The 14-3-3 proteins constitute a family of abundant, highly conserved and broadly expressed acidic polypeptides. One member of this family, the 14-3-3σ isoform {sigma}, is expressed only in epithelial cells and is frequently down-regulated in a variety of human cancers and plays a role in the cellular response to DNA damage. The 14-3-3s generally form heterodimers with other family members, but 14-3-3σ preferentially forms homodimers in cells. Three amino acids that are completely conserved in all other 14-3-3s, are not present in 14-3-3σ. These amino acids unique to 14-3-3σ confer a second ligand-binding site involved in 14-3-3σ-specific ligand discrimination.

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