Discovery and functional characterization of cytotoxic cells in planarians

Research into functional cellular immune activity in invertebrates, specifically phagocytosis and cytotoxicity, remains an evolving field with ample room for exploration. Phagocytosis has garnered relatively more attention, while cytotoxicity, a fundamental aspect of immune defense, has been understudied or indirectly addressed in some invertebrate species.

Planarians, renowned for their extraordinary regenerative abilities, thus have predominantly been investigated in the context of regeneration. They exhibit immune-related functions, including evidence of phagocytic activity directed towards bacteria and cellular components, and some hints of cytotoxic activity. This study delves into the cellular immune capabilities of planarians, presenting the first evidence of cellular cytotoxic activity within the planarian species, by means of flow cytometry-based functional immune assays. Furthermore, this research uses live visualizations that capture the dynamic interactions of cytotoxic cells with their targets using fluorescence labeling of isolated cells and confocal microscopy. In addition, it sheds light on their ability to phagocytose entire cells. Remarkably, using functional immune assays of isolated cells of syngeneic, allogeneic, and xenogeneic species, we reveal that these immune responses do not exhibit specific cellular activity during encounters between allogeneic and xenogeneic cells, suggesting a lack of specificity in recognizing incompatible cells. This observation, combined with additional discoveries, strengthened our hypothesis that planarian cytotoxic cells belong to the level-I cytotoxicity category. They lack the ability to specifically identify incompatible cells and instead engage in non-specific killing once activated.

Furthermore, we successfully isolated a granular cell population enriched in cytotoxic cells. This enriched population demonstrates enhanced target cell-killing capabilities and exhibits upregulation of genes associated with cytotoxic activity. Several key characteristics align these planarian cytotoxic cells with neutrophil-like cells. These findings expand our comprehension of cytotoxicity in invertebrates and shed some light on the evolution of cellular immune lineages. Notably, these findings represent the first comprehensive characterization of early-stage cytotoxicity in this class of organisms.