Figure 4

Changes in the expression of Rho-GTPase genes (RhoA, Rac1 and Cdc42) induced by extracellular vesicles secreted by trypomastigotes of T. cruzi. (A) An overexpression of RhoA and Rac1 was observed 15 min post-incubation of Vero cells with EV, while the expression of Cdc42 was downregulated. After this time- point, a downregulation in the expression of the 3 analyzed genes appeared at 30 min, being more evident 4 h post-interaction. The incubation of cells with trypomastigotes revealed an overexpression of the 3 genes both 15 and 30 min post-interaction, while the same behavior of EV in downregulating the expression of the 3 genes was observed 24 h post-interaction. Gene expression was calculated as fold induction caused by the respective treatment/control as compared with the expression of GAPDH. Data are presented as shown above, with mean ± S.D. (n = 3). Differences to this value were analyzed by one-way ANOVA using the Tukey’s post correction as rank tests (asterisks), where * p < 0.05; ** p < 0.01; **** p < 0.0001, comparing within the same time stamp the trypomastigotes (TC, orange) and the EV controls (blue). Comparison of each time point against the 15 min time are represented as †, for the TC groups, and ‡ for the EV groups. Expression levels of SRGAP3, an activating protein of Rho-GTPases, in cells incubated with EV of trypomastigotes of T. cruzi for 4 h, derived from the mRNA levels (B) and on protein levels (C). Expression was calculated as fold induction caused by the respective treatment/control as compared with the expression of GAPDH. Data are presented as shown above, with mean ± S.D. (n = 3). Differences to this value were analyzed by one sample Tukey multiple comparison test signed rank tests (asterisks), where *** p < 0.001.