Reassessment of endothelin receptor A expression in normal and neoplastic human tissues using the novel rabbit monoclonal antibody UMB-8

The endothelins ET-1, ET-2 and ET-3 are a family of peptides, which exert their actions via two G protein-coupled receptor subtypes, ETA and ETB. ET-1 is a potent vasoconstrictor and is involved in the development of different cardiovascular and renal disorders. Additionally, ET-1 and the ETA receptor have been shown to be important mediators of cancer growth and metastasis. We have extensively characterized the novel monoclonal rabbit anti-ETA antibody UMB-8 using transfected cells as well as mouse, rat and human tissues. UMB-8 was then tested in a large series of formalin-fixed and paraffin-embedded human normal and neoplastic tissue specimens. Specificity of UMB-8 was demonstrated by detection of a broad band migrating at 70-80 kDa in Western blot analyses of ETA-transfected HEK-293 cells and of different mouse tissues and by agonist-dependent translocation of the immunosignal from the surface of ETA-transfected cells into the cytosol. In tissue samples, UMB-8 yielded an efficient immunostaining of distinct cell populations with a predominance of plasma membrane staining, which was abolished after preadsorption of the antibody with its immunizing peptide. In normal tissue, ETA was present in the heart, blood vessels, gut and kidneys. ETA was also detected with a hitherto unappreciated high prevalence in all types of sarcomas and in glioblastomas, but also in various epithelial tumor entities and in tumor stroma. All in all, UMB-8 may prove of great value in the identification of ETA-expressing tumors during routine histopathological examinations. (C) 2015 Elsevier Inc. All rights reserved.