Crotoxin modulates Encephalitozoon cuniculi-infected macrophages toward the M1 microbicidal profile

Crotoxin (CTX), a bioactive extract from the snake Crotalus durissus terrificus, has antibacterial, antitumor, and anti-inflammatory properties. Microsporidia are opportunistic, obligate intracellular fungi that infect vertebrates and invertebrates and are highly resistant to conventional drugs. They can also subvert the microbicidal activity of M1 macrophages to an M2 profile, which is more favorable for the pathogen. Thus, in this study, we evaluated the effects of CTX on the viability of spores of the microsporidium Encephalitozoon cuniculi, as well as on the microbicidal activity of macrophages in vitro. E. cuniculi spores were treated with two concentrations of CTX (2.4 and 4.8 μg/mL) and cultivated in RK-13 cells for viability analysis. Additionally, peritoneal adherent cells (APerC), obtained from peritoneal washes of BALB/c mice, were infected with spores of E. cuniculi for 1 h and treated with CTX for 3 h. The profile of macrophages, cytokine production, viability of macrophages, and proliferative capacity of spores were subsequently evaluated. Treatment of E. cuniculi spores with CTX had no fungicidal or fungistatic effects. Compared to the macrophages in the control group, macrophages infected with E. cuniculi and treated with 2.4 μg/mL CTX presented an increase in the M1 profile, more necrosis, and greater production of the cytokines TNF-α and IL-6, and the spores obtained from these macrophages presented a reduction in proliferative capacity. These results indicated that CTX modulated the M1 profile of macrophages infected with E. cuniculi, resulting in greater production of proinflammatory cytokines and stronger microbicidal activity.