Receptor binding and biologic activity of synthetic ET-1 peptides in the retinal pericyte

The purpose of this study was to examine the effect of synthetic endothelin (ET)-1 peptides with antigenic potential for binding and biologic activity using an in vitro model of microvascular pericytes.

All possible sequential hexapeptide fragments of endothelins -1, -2, and -3 were synthesized on polyethylene rods and tested for reactivity with antibodies for ET-1 and ET-3. The most highly antigenic peptide ET-1[3-8] and the least antigenic ET-1[1-6], which gave low reactivity in the enzyme-linked immunosorbent assay (ELISA), were synthesized. The C-terminal hexapeptide ET-1[16-21], which has been reported as an ETB receptor agonist but which gave no reactivity in the ELISA, also was investigated. The synthesized ET analogues were tested for receptor binding, inositol phosphate generation, and mitogenesis in bovine retinal pericytes.

ET-1[16-21] partially inhibited both [125I]-ET-3 binding at high concentrations, as did ET-1[1-6]. In contrast, ET-1[3-8] displaced labeled ET-1 binding but not labeled ET-3 binding. None of the peptides had any significant mitogenic or second-messenger responses when compared to those elicited by the full ET-1 molecule.

The result of the current work shows that the sequence, ET[3-8], is involved in isopeptide-specific binding to ETA receptors, but it suggests that other regions of the molecule are necessary for full bioactivity in microvascular pericytes.