Fig 2.

(A-F) Parp1^-/- primary Mφ exhibit decreased cytokines’ expression in response to TEv stimulation. Supernatants of Raw 264.7 Mφ incubated with media only or T. cruzi were used to purify NEv_RAW and TEv_RAW, respectively. Primary bone-marrow Mφ (WT or Parp1^-/-) were incubated with Ev (± 20 ng/mL IFN-γ) for 3 h and 48 h. (A-C) Real time RT-qPCR analysis of mRNA levels for cytokine encoding genes at 3 h post-incubation. (D-F) ELISA determination of cytokines release at 48 h post-incubation. Primary Mφ incubated with T. cruzi (cell: parasite ratio, 1:3) and IFN-γ were used as controls. (G-I) Immune characteristics of Ev produced during the development of CD (± PARP1 inhibitor). B6129S/J WT mice (n = 10 per group) were infected with T. cruzi (10,000 parasites per mouse). Plasma of normal and chronically infected (150 days post-infection) mice were used to isolate NEv_mice and TEv_mice^ch, respectively. Next, Mφ were incubated with murine plasma Ev (± 20 ng/ml IFN-γ and 5 μM iniparib) for 48 h, and an ELISA was performed to determine the TNF-α (G), IL-6 (H), and IL-1β (I) release in supernatants. Data are representative of ≥ 2 independent experiments (two biological replicates per treatment and triplicate observations per sample for RT-qPCR analysis; and three biological replicates per treatment and duplicate observations per sample for ELISA). Data are plotted as mean value ± SD. Significance is annotated as ⁺ NEv vs. TEv, * TE_V vs. TEv+IFN-γ, and ⁱ WT.Tc vs. Parp1^-/-.Tc and or effect of iniparib on TEv+IFN-γ; and p values of ≤ 0.05, ≤ 0.01, and ≤ 0.001 are presented by one, two, and three symbol characters, respectively. Horizontal bars show the compared groups.