In vitro spontaneous cytotoxic activity against mammalian target cells by the hemocytes of the solitary ascidian, Ciona intestinalis

Abstract

Blood cell-mediated cytotoxic activity against mammalian target cells by the hemocytes of the solitary ascidian Ciona intestinalis was investigated in vitro by fluorochromasia. Salt-conditioned target cells were labeled with carboxyfluorescein diacetate and challenged with mixed and separated hemocytes. The assay provided optimal conditions for the functioning of the effector hemocytes while maintaining low background leakage from the target cells. Comparison of different hemocyte populations, separated by density gradient centrifugation, revealed that only cell bands containing the phagocytic and nonphagocytic amoebocytes exhibited cytotoxicity. Experiments to characterize cytolysis demonstrated that activity increased with the effector to target cell ratio, occurred within 15 min, and was maximal at an incubation temperature of 20 degrees C. Both human (K562) and mouse [YAC-1, P815, WEHI (3B) and L929] target cell lines were killed by the ascidian effector hemocytes. This paper demonstrates a population of nonspecific cytotoxic effector cells in the blood of C. intestinalis that are able to spontaneously kill a range of mammalian targets in vitro.