Differentiation of blood cell lines in the colonial ascidian Botryllus schlosseri.

Ascidian blood contains a wide variety of morphologically different hemocytes which, in spite of numerous studies, are still matter of debate due to the persisting doubts and uncertainties about their differentiation pathways, mutual relationships and functions. Our interest in the role of blood cells in defence strategies of the colonial ascidian Botryllus schlosseri has led us to study and characterize two differentiation lines of hemocytes involved in two defence reactions, namely phagocytosis which preserves the organism from invading microorganisms and histocompatibility which prevents the fusion of genetically remote colonies. Three hemocyte types take part in phagocytosis, i.e. hyaline amoebocytes, macrophage-like cells and signet-ring cells: all of them share a common enzymatic pattern rich in hydrolytic enzymes. Hyaline amoebocytes can rapidly attach and ingest test particles; phagosomes inside macrophage-like cells and signet-ring cells appear after 30 min of incubation with test particles. These data suggest that they may represent different functional aspects of a single cell-type. This hypothesis is supported by the study of hemocyte frequencies in the weekly life cycle of Botryllus: during the resorption of the old blastogenetic generation, characterized by intense phagocytic activity, a significant decrease in the number of hyaline amoebocytes occurs together with a significant increase in macrophage-like cells. Histocompatibility in Botryllus manifests itself as rejection reaction between contacting colonies and is characterized by the appearance of dark-brown necrotic masses at the borderline. In this reaction the so called morula cells (MC) play an important role, as they accumulate at the apices of contacting ampullae, cross the ampullar epithelium and migrate into the tunic where they finally release the content of their vacuoles and degenerate. MC have granular amoebocytes as their precursors and contain polyphenolic substance and phenoloxidase (PO) inside their vacuoles: in our hypothesis PO, by transforming plyphenol substrata into quinones, is the enzyme responsible of the localized cytotoxicity during the rejection reaction. We have demonstrated that MC degranulate and release the vacuolar content when incubated with incompatible blood plasma (BP) and an increase in cell mortality is observed when hemocytes are incubated for 60 min with incompatible BP.