Localisation of apelin-12, a ligand for the orphan G-protein coupled with APJ receptor, to human cardiovascular tissue Matthias Kleinz, Rhoda Kuc, Janet Maguire & Anthony Davenport, Clinical Pharmacology Unit, University of Cambridge, Level 6, Centre for Clinical Investigation, Box 110 Addenbrooke's Hospital, Cambridge, CB2 2QQ, UK.

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Apelin, a 36 amino acid peptide recently isolated from bovine stomach, is the endogenous ligand of the orphan G-protein coupled receptor APJ (Tatemoto et al., 1998). We have pre-viously demonstrated the presence of high affinity [¹²⁵I]apelin-13 binding sites in human heart, aorta, coronary artery and sa-phenous vein and identified p[Glu] apelin-13 as a potent vaso-constrictor of human saphenous vein (Katugampola et al., 2001). Our aim was to determine the distribution of apelin-12-like immunoreactivity in human tissues using an antibody raised against the C-terminal dodecapeptide of apelin.

For immunocytochemistry, human tissues were obtained with ethical approval. Left ventricle and atria were from patients transplanted for ischaemic heart disease (n=8), cardiomyopathies (n=6), cystic fibrosis (n=3), or from donor hearts for which there was no suitable recipient (n=3). Sections of histologically normal kidney (n=5) and lung (n=5) were from patients undergoing nephrectomy and lobectomy respectively for non-obstructive carcinomas. Adrenal was from one patient undergoing adrenalectomy for phaeochromocytoma.Cryostat sections (10µm) of tissues were fixed in ice-cold acetone for 10 min. Sections were blocked with 5% swine serum in phosphate-buffered saline (PBS) for 1h at 23°C and incubated with an anti-apelin-12 polyclonal antibody (Phoenix Pharmaceuticals, CA. USA) at a dilution of 1:500, at 4°C for 72h, in PBS containing 0.01% tween-20 and 1% swine serum. Specific staining was detected using the peroxidase anti-peroxidase method. Adjacent sections were incubated with non-immune serum as a negative control.

Fig. 1 Apelin-LI localised to human intramyocardial (A) and adrenal plexus (B) arteries. Scale bar = 200µm.

Intense apelin-12-like immunoreactivity (apelin-LI) was localised to the endothelium of intramyocardial (Fig. 1), intrarenal and small pulmonary blood vessels and to endocardial endothelial cells lining the atria. Little or no staining was observed in smooth muscle, cardiomyocytes or epithelial cells in lung or kidney. In adrenal gland, apelin-LI was confined to endothelial cells of surrounding arteries, small resistance arteries within the capsular plexus (Fig. 1) and the central vein. Staining was not detected in secretory cells of the adrenal medulla and cortex.The presence of apelin-LI in vascular endothelial cells sug-gests that in the human cardiovascular system apelin may act as a vasoactive mediator that elicits its function by activation of receptors present on the underlying vascular smooth muscle and myocardium.

MK supported by Isaac Newton and Cambridge European Trusts.

Katugampola, S.D., Maguire, J.J., Matthewson, S.R. & Davenport, A.P. (2001). Br. J. Pharmacol., 132, 1255-1260.
Tatemoto, K., Hosoya, M., Habata, Y., et al., (1998). Biochem. Biophys. Res. Commun. 251, 471-476.