RGD-dependent mechanisms in the endoneurial phagocyte response and axonal regeneration in the nervous system of the snail Lymnaea stagnalis

Abstract

Activation of phagocytic cells in the injury zone is a crucial step in the regeneration of peripheral axons. Many aspects of the mechanisms underlying the recruitment of active phagocytes remain, however, unclear. Notably, our understanding of the interactions between injury, extracellular matrix (ECM) degradation and phagocyte activation is limited. Most animal cell types, phagocytes included, interact with proteins of the ECM through one or more members of the integrin family, transmembrane cell adhesion receptors that typically bind their ligands through short linear amino acid sequences. This study focused on the role of one of the most common of such integrin recognition sequences, the Arg-Gly-Asp (RGD) motif in the recruitment and activation of endoneurial phagocytes in the injury response of the nervous system of the pond snail Lymnaea stagnalis. Like the mammalian nervous system, the Lymnaea nervous system responds to injury with recruitment and activation of endoneurial phagocytes (i.e. phagocytes residing in Lymnaea's nerves), a process involving substantial changes in the morphology, motility and adhesion status of these cells. Using synthetic water-soluble RGD-peptides, we investigated the relevance of RGD-dependent mechanisms in the activation of endoneurial phagocytes and injury response of the organ-cultured nervous system of Lymnaea. Our results show that RGD-peptides modulate various aspects of phagocyte activation (i.e. spreading response, particle engulfment, oxidative burst) in vitro and in situ and significantly affect nerve regeneration in this model system. Surprisingly, while linear RGD-analogues suppressed both phagocyte activation and axonal regeneration, a circularized RGD-peptide analogue modulated these parameters in a concentration-dependent, biphasic manner. Collectively, these results emphasize the significance of RGD-dependent mechanisms in the regenerative response of the Lymnaea nervous system and implicate regulation of the cellular immune response as one of the factors in this context.