Synthetic complementary peptides inhibit a neutrophil chemoattractant found in the alkali-injured cornea

Purpose. We have previously presented evidence that the neutrophil chemoatt ractant, N-acetyl-proline-glycine-proline (N-acetyl-PGP), triggers the init ial polymorphonuclear leukocyte (PMN) invasion into the alkali-injured eye. In this study, sense-antisense methodology was used to develop novel compl ementary peptides to be potential inhibitors of N-acetyl-PGP. Methods. The polarization assay was used to measure the potential chemotactic response o f PMNs to synthetic N-acetyl-PGP, the ultrafiltered tripeptide chemoattract ants obtained from alkali-degraded rabbit corneas, or leukotriene B-4 (LTB4 ). Inhibition was expressed as the peptide concentration producing 50% inhi bition (ID50) of polarization. Five complementary peptides were tested as p otential inhibitors of N-acetyl-PGP: arginine-threonine-arginine (RTR), RTR -glycine-glycine (RTRGG), RTR dimer, RTR tetramer, and alanine-serine-alani ne (ASA) tetramer. In addition, the RTR tetramer and both monomeric peptide s (RTR and RTRGG) were separately tested for inhibition of the ultrafiltere d tripeptide chemoattractants or LTB4. Results, The complementary RTR tetra meric peptide was a powerful antagonist of N-acetyl-PGP-induced PMN polariz ation (ID50 of 200 nM). The RTR dimer was much Less potent (ID50 of 105 mu M). Both monomeric peptides, RTR and RTRGG, were only antagonistic at milli molar concentrations. The ASA tetramer showed no capacity to inhibit N-acet yl-PGP. The RTR tetramer also inhibited PMN activation by the ultrafiltered tripeptide chemoattractants (ID50 of 30 mu M) but had no effect on LTB4. C onclusions. A complementary peptide (RTR) was designed which is an effectiv e inhibitor of the neutrophil chemoattractant, N-acetyl-PGP. The potency of the RTR complementary peptide is dramatically enhanced by tetramerization. Inhibition of N-acetyl-PGP by complementary peptides offers great promise for control of the inflammatory response in the alkali-injured eye.