Localization of endothelin peptides in human kidney

Abstract

We investigated the synthesis and localization of endothelin isoforms in the human kidney using the reverse-transcriptase polymerase chain reaction (RT-PCR) and immunocytochemistry. PCR products corresponding to the expected size for mRNA encoding ET-1, ET-2 and ET-3 were found in homogenates of renal medulla, cortex and vessels from each of five individuals. Using four rabbit polyclonal antibodies to assess the distribution of mature ET, Big ET-1, Big ET-2 and Big ET-3 immunoreactivity in the human kidney, mature IR ET localized to the cytoplasm of endothelial cells lining intra-renal blood vessels including interlobular and arcuate arteries, arterioles and adjacent arcuate veins, all of which showed strongly positive staining. IR Big ET-1 co-localized with the mature peptide. No specific staining was detected within these anatomical regions when pre-immune sera were substituted or primary antibody omitted. Mature IR ET also localized to the cytoplasm of endothelial cells within the glomerulus. Other capillary endothelial cells did not stain, and other structures stained only faintly by comparison. IR Big ET-2 and Big ET-3 could not be detected. These results show that human kidney contains mRNA encoding all three peptide isoforms, but only mature ET and Big ET-1 peptides could be detected by immunocytochemical staining. This provides further evidence that ET-1 may function as a renal peptide in humans, as it is locally synthesized within the kidney.