IFN-inducible p47 GTPases display differential responses to Schistosoma japonicum acute infection

Abstract

Interferon gamma induced GTPase (IGTP) (also named Irgm3) and interferon gamma inducible protein 47 (IRG-47) (also named Irgd) are interferon (IFN)-inducible p47 GTPases that have been shown to regulate host resistance to intracellular pathogens. Little knowledge has been known about the role of p47 GTPases in host responses against extracellular pathogens. To investigate possible roles of IGTP and IRG-47 in the course of Schistosoma japonicum infection, IGTP and IRG-47 knockout and wild-type (WT) mice were challenged with cercariae of S. japonicum, and host responses were analyzed. At the acute stage of S. japonicum infection, mice that lacked IGTP displayed similar parasite burden and pathological damage to WT mice. Importantly, S. japonicum-infected IRG-47-deficient mice, in contrast to IGTP-deficient mice and WT mice, showed significantly reduced worms and lower egg-burden, but intense granulomatous reaction evoked by schistosome eggs in peripheral parts of liver lobes. In addition, upregulation of inflammation-related gene expression was observed in the spleen of IRG-47-deficient mice using oligonucleotide microarrays, in which multiple pathways of cytokine-cytokine receptor interaction, T-cell receptor signaling, complement, coagulation cascades and cell adhesion molecules were highlighted. Taken together, these data suggest that IGTP and IRG-47 might have distinct features that were differentially required for resistance to S. japonicum.

Figure 1

Liver pathology with HE staining in 6-week-S. japonicum-infected IGTP^−/−, IRG-47^−/− and WT mice, respectively. (a–c) A representative portrait of the liver portal area from IGTP^−/−, IRG-47^−/− and WT mice at a magnification of ×100 (scale bar=400 μm). (d–f) A representative picture of granuloma with a single egg from IGTP^−/−, IRG-47^−/− and WT mice at a magnification of ×400 (scale bar=100 μm).

Figure 2

The cellularity in liver granuloma with a single egg in 6-week-S. japonicum-infected IGTP^−/−, IRG-47^−/− and WT mice, respectively. Eosinophils, lymphocytes, plasma cells and the total cells were counted in each granuloma. At least 10 granulomas were counted in each section of mice. Asterisks indicate significant differences (*P<0.05, **P<0.01, compared with WT mice).

Figure 3

The amounts of Th1/Th2 cytokines in the sera from 6-week-S. japonicum-infected IGTP^−/−, IRG-47^−/− and WT mice were assayed by Bio-Plex technology. The symbol ‘▴' presents the absolute amount of each sample. Asterisks indicate significant differences (*P<0.05, **P<0.01).

Figure 4

Functional classification of 26 selected, differentially expressed genes from microarray data of spleen cells. Genes were divided into immunologically relevant functional families. Each colored box represents the normalized expression level of a given gene in a particular experimental condition and is colored according to the ratio (log₂(signal intensity of IGTP^−/− or IRG-47^−/−)/signal intensity of WT) of a specific gene-expression level in IGTP^−/− or IRG-47^−/− mice to that in WT mice.

Figure 5

Significant pathways with differentially changed genes in IGTP^−/− and IRG-47^−/− mice compared with WT mice. The bioinformatics analysis was carried out using GeneSpring version 7.0 software. Differentially changed genes were placed into Kyoto Encyclopedia of Genes and Genomes pathways database to search the relevant pathways. Significant pathways associated with immune and inflammatory response in IRG-47^−/− mice focused on cell adhesion, cytokine–cytokine receptor interaction, T–cell receptor signaling, complement and coagulation.

Figure 6

The relative mRNA levels of ccl2, il12, il10 and three members of p47 GTPase family, igtp, lrg47 and irg47, at 6-week-S. japonicum-infected IGTP^−/−, IRG-47^−/− and WT mice, were assayed by real-time quantitative PCR. The ratio of target gene to internal control was calculated to represent the relative expression level. (a) The relative mRNA levels of ccl2, il12 and il10 indicated that the expressions of ccl2, il12 and il10 in IRG-47^−/− mice were higher than those in IGTP^−/− and WT mice. Especially, ccl2 expression in IRG-47^−/− mice presented a significant increase (*P<0.05). (b) lrg47 gene was significantly upregulated in both IGTP^−/− and IRG-47^−/− mice, compared with WT group (*P<0.05).