Modulation of neutrophil phospholipase C activity and cyclic AMP levels by fMLP-OMe analogues

The N-formyl- l-methionyl- l-leucyl- l-phenylalanine (fMLP)-OMe ( 1) analogues for-Thp-Leu-Ain-OMe ( 2), for-Thp-Leu-Phe-OMe ( 3), for-Met-Leu-Ain-OMe ( 4), for-Met-Δ zLeu-Phe-OMe ( 5), for-Met-Lys-Phe-For-Met-Lys-Phe ( 6), for-Met-Leu-Pheol-COMe ( 7), and for-Nle-Leu-Phe-OMe ( 8) have been studied. Some of these have been found selective towards the activation of different biological responses of human neutrophils. In particular, peptides 2 and 3, which evoke only chemotaxis, are ineffective in enhancing inositol phosphate, as well as cyclic AMP (cAMP) levels. On the contrary, analogues 5 and 7, which induce superoxide anion production and degranulation, but not chemotaxis, significantly increase the levels of the two intracellular messengers, as is the case of the full agonists 1 and 6. The Ca 2+ ionophore A23187 also activates phospholipase C (PLC) and increases the nucleotide levels; when tested in combination with peptide 1 or 5, a supra-additive enhancement of cAMP concentration is obtained. The PLC blocker, U-73122, inhibits the formylpeptide-induced inositol phosphate formation, as well as cAMP increase. Moreover, this drug drastically reduces superoxide anion release triggered by 1 or 5, whereas it inhibits to a much lesser extent neutrophil chemotaxis induced by 1 or 2. Our results suggest that: (i) PLC stimulation is involved in cAMP enhancement by formylpeptides; (ii) the activation of PLC by formylpeptides, in conditions of increased Ca 2+ influx, induces a supra-additive enhancement of the nucleotide; (iii) the inability of pure chemoattractants to significantly alter the PLC activity or cAMP level, differently from full agonists or peptides specific in inducing superoxide anion release, appears as a general property. Thus, the activation of neutrophil PLC seems essential for superoxide anion release, but less involved in the chemotactic response.