During acute infection with Trypanosoma cruzi, the causative agent of Chagas' disease, the thymus undergoes intense atrophy followed by a premature escape of CD4 ؉ CD8 ؉ immature cortical thymocytes. Here we report a pivotal role for the endogenous lectin galectin-3 in accelerating death of thymocytes and migration of these cells away from the thymus after T. cruzi infection. We observed a pronounced increase in galectin-3 expression that paralleled the extensive depletion of CD4 ؉ CD8 ؉ immature thymocytes after infection. In vitro, recombinant galectin-3 induced increased levels of death in cortical immature thymocytes. Consistent with the role of galectin-3 in promoting cell death, thymuses from gal-3 ؊/؊ mice did not show cortical thymocyte depletion after parasite infection in vivo. In addition, galectin-3 accelerated laminin-driven CD4 ؉ CD8 ؉ thymocyte migration in vitro and in vivo induced exportation of CD4 ؉ CD8 ؉ cells from the thymus to the peripheral compartment. Our findings provide evidence of a novel role for galectin-3 in the regulation of thymus physiology and identify a potential mechanism based on proteinglycan interactions in thymic atrophy associated with acute T. cruzi infection.