Interleukin-25-mediated resistance against intestinal trematodes does not depend on the generation of Th2 responses

Abstract

Background: The cytokine interleukin-25 (IL-25) is recognized as the most relevant initiator of protective T helper 2 (Th2) responses in intestinal helminth infections. This cytokine induces resistance against several species of intestinal helminths, including the trematode Echinostoma caproni. E. caproni has been extensively used as an experimental model to study the factors determining resistance to intestinal infections. In the study reported here, we assessed the role of IL-25 in the generation of resistance in mice infected with E. caproni.

Methods: The factors that determine the production of IL-25 in mice experimentally infected with E. caproni were determined, as were the consequences of IL-25 production in terms of polarization of the immune response and resistance to infection.

Results: Our results show that the role of IL-25 in the polarization of the immune response differs between the primary and secondary immune responses. IL-25 is required for the development of a Th2 phenotype in primary E. caproni infections, but it can also promote the differentiation to Th2 memory cell subsets that enhance type-2 immunity in memory responses. However, the development of Th2 responses does not induce resistance to infection. The Th2 phenotype does not elicit resistance, and IL-25 is responsible for the resistance regardless of its type-2 cytokine activity and activation of signal transducer and activator of transcription (STAT6). Alternative activation of macrophages induced by IL-25 can be implicated in the resistance to infection.

Conclusions: In contrast to primary infection, secondary infection elicits a type-2 immune response even in the absence of IL-25 expression. Despite the development of a type-2 response, mice are susceptible to secondary infection associated with the lack of IL-25. Resistance to infection is due to the production of IL-25, which acts autonomously from Th2 response in terms of parasite clearance.

Keywords: Echinostoma caproni; Interleuquin-25; Intestinal helminth; Resistance; Th2; Trematoda.

Fig. 1

Blocking of interleukin-25 (IL-25) in challenge infections with Echinostoma caproni reverts resistance to infection despite the development of a T-helper cell (Th2) response. A Worm recovery in IL-25 monoclonal antibody (mα-IL-25)-treated ICR mice and non-treated mice after a challenge infection with E. caproni. B Expression of cytokine mRNA in the intestinal tissue of mα-IL-25-treated ICR mice and non-treated mice after a challenge infection with E. caproni. The relative quantities (RQ) of cytokine genes are shown after normalization with β-actin and standardization of the relative amount against the day 0 sample. Vertical bars represent the standard deviation (SD). A Asterisk indicates significant difference from non-mα-IL-25-treated mice, B lowercase letters (a, b) above bars indicate either significant differences with respect to naïve mice controls (a) or significant differences between groups (b), both at p < 0.05

Fig. 2

Blocking of IL-25 induced alternative activation of macrophages after challenge infection. A Pattern of macrophage activation is different in primary and secondary infections analyzed according to the gene expression of marker mRNA of both classical (ArgII and iNOS) and alternative (ArgI and Ym-1) macrophage activation in the intestinal tissue of mα-IL-25-treated mice and non-treated mice after a challenge infection with E. caproni. B Expression of resistin-like molecule beta (RELM-β) mRNA in the intestinal tissue of mα-IL-25-treated ICR mice and non-treated mice after a challenge infection with E. caproni. The RQ of cytokine genes are shown after normalization with β-actin and standardization of the relative amount against the day 0 sample. Vertical bars represent the SD. Lowercase letters (a, b) above bars indicate either significant differences with respect to naïve mice controls (a) or significant differences between groups (b), at p  < 0.05

Fig. 3

Interleukin-25 (IL-25) gene expression returned to baseline levels 10 weeks after pharmacological cure of a primary infection. Expression of IL-25 mRNA in the intestinal tissue mice after curation with praziquantel of a primary infection with E. caproni. The RQ of cytokine genes are shown after normalization with β-actin and standardization of the relative amount against the day 0 sample. Vertical bars represent the SD

Fig. 4

Recovery of baseline expression of IL-25 mRNA after healing of the primary infection reverted the resistance against challenge infection together with a Th2 response. A Worm recovery of a primary infection in naïve mice and in that of a challenge infection in mice in which the basal levels of mRNA expression were recovered after the cure of a primary infection. B Expression of cytokine mRNA in the intestinal tissue of both groups of mice at 2 weeks after the primary and secondary infection (wppi and wpsi, respectively). The RQ of cytokine genes are shown after normalization with β-actin and standardization of the relative amount against the day 0 sample. Vertical bars represent the SD. Lowercase letters (a, b) above bars indicate either significant differences with respect to naïve mice controls (a) or significant differences between groups (b), at p  < 0.05

Fig. 5

Treatment of mice with either mα-IL-4Rα or rIL-13Rα2 abrogates Th2 response and alters the pattern of macrophage activation despite the presence of IL-25. A Expression of cytokine mRNA in the intestinal tissue IL-25-treated-mice that were also treated with monoclonal anti-mouse IL-4Rα (mα-IL-4Rα) or recombinant IL-13Rα2 (rIL-13Rα2) at 2 weeks post-primary infection (wppi) with E. caproni; B Pattern of macrophage activation analyzed by the expression of marker mRNA of both classical (ArgII and iNOS) and alternative (ArgI and Ym-1) macrophage activation in the intestinal tissue of IL-25-treated-mice that were also treated with mα-IL-4Rα or rIL-13Rα2 at 2 wppi with E. caproni. The RQ of cytokine genes are shown after normalization with β-actin and standardization of the relative amount against the day 0 sample. Vertical bars represent the SD. Lowercase letters (a, b) above bars indicate either significant differences with respect to naïve mice controls (a) or significant differences between groups (b), at p  < 0.05

Fig. 6

Pharmacological curation of an E. caproni primary infection exacerbates the expression of IL-13Ra2. A Schematic representation of the experimental protocol. B Levels of mRNA expression of IL-13Ra2 in the intestinal tissue of mice primarily infected at 4 wpi, at 2 weeks post-treatment with praziquantel and at 2 wpsi. The relative quantities (RQ) of cytokine genes are shown after normalization with β-actin and standardization of the relative amount against day 0 sample. Vertical bars represent the standard deviation. Lowercase letters (a, b) above bars indicate either significant differences with respect to naïve mice controls (a) or significant differences between groups (b), at p  < 0.05