Morula cells and non-fusion reaction in the compound ascidian Botryllus schlosseri

Morula cells (MC) are a common haemocyte-type in the compound ascidian Botryllus schlosseri, their frequency ranging between 40 and 60% of the circulating blood cells. They are the effector cells of the non-fusion reaction, characterised by the appearance of necrotic foci along the contact border, which occurs when genetically incompatible colonies contact each other. We previously demonstrated that, in the course of this reaction, MC acquire immunopositivity to anti-cytokine (IL-1-a and TNF-a) antibodies, degranulate and release the enzyme phenoloxidase which is responsible of the cytotoxicity observed both in vitro (when haemocytes are incubated with blood plasma (BP) from incompatible colonies) and in vivo (non-fusion reaction). Subsequently, MC leave the facing marginal ampullae (sausage-like, blind endings of the colonial marginal vessels to reach the tunic, apparently attracted by soluble, diffusing factors, where they degenerate and contribute to the formation of the cytotoxic spots. In the present work, we focussed on the chemotactic recruitment of MC in the course of the nonfusion reaction. As a first approach, we studied the distribution of MC inside the facing marginal ampullae of both contacting colonies (either non-fusible or fusible) and solitary ones. Results clearly indicates a significantly higher concentration of MC inside facing marginal ampullae of incompatible colonies with respect to compatible colonies; the latter concentration is, however, significantly higher than that inside ampullae from solitary colonies. In addition, we used Transwell chambers to evaluate whether incompatible BP has chemotactic properties. We put haemocyte suspensions in filtered sea water (FSW) in the upper wells and BP from either incompatible or compatible colonies in the lower wells. We observed a significant increase migration of haemocytes, and of MC in particular, in the presence of incompatible BP. This migration was significantly decreased by the addition of anti-cytokine(IL-1-a, TNF-a or IL-8) antibodies, suggesting that molecules molecules recognised by these antibodies can be responsible of the chemotaxis observed.