Genetic control of severe egg-induced immunopathology and IL-17 production in murine schistosomiasis

Abstract

Infection with the trematode parasite Schistosoma mansoni results in a distinct heterogeneity of disease severity, both in humans and in an experimental mouse model. Severe disease is characterized by pronounced hepatic egg-induced granulomatous inflammation in a proinflammatory cytokine environment, whereas mild disease corresponds with reduced hepatic inflammation in a Th2 skewed cytokine environment. This marked heterogeneity indicates that genetic differences play a significant role in disease development, yet little is known about the genetic basis of dissimilar immunopathology. To investigate the role of genetic susceptibility in murine schistosomiasis, quantitative trait loci analysis was performed on F(2) progeny derived from SJL/J and C57BL/6 mice, which develop severe and mild pathology, respectively. In this study, we show that severe liver pathology in F(2) mice 7 wk after infection significantly correlated with an increase in the production of the proinflammatory cytokines IL-17, IFN-gamma, and TNF-alpha by schistosome egg Ag-stimulated mesenteric lymph node cells. Quantitative trait loci analysis identified several genetic intervals controlling immunopathology as well as IL-17 and IFN-gamma production. Egg granuloma size exhibited significant linkage to two loci, D4Mit203 and D17Mit82, both of which were inherited in a BL/6 dominant manner. Furthermore, a significant reduction of hepatic granulomatous inflammation and IL-17 production in interval-specific congenic mice demonstrated that the two identified genetic loci have a decisive effect on the development of immunopathology in murine schistosomiasis.

FIGURE 1

Granuloma size and IL-17 production by SEA-stimulated MLN cells from SJL, BL/6, F₁, and F₂ mice. A, Granulomas were measured in liver sections obtained from SJL (n = 19), BL/6 (n = 20), F₁ (n = 10) and F₂ mice (n = 150) 7 wk after infection, as described in Materials and Methods. A minimum of 20 granulomas were measured per mouse. Each dot represents an individual mouse. B, IL-17 production by SEA-stimulated MLN cells from these mice was measured in 48-h culture supernatants by ELISA, as described in Materials and Methods. Each dot represents the mean cytokine level of triplicate determinations per mouse. Black horizontal lines represent the mean of the group.

FIGURE 2

Linear regression analysis of mean granuloma size vs cytokine production for individual F₂ mice. IL-17 (A), IFN-γ (B), and TNF-α (C) production from MLN cells significantly correlate with granuloma size in F₂ mice. There was no correlation between IL-4 (D), IL-5 (E), and IL-10 (F) production and granuloma size in these mice. Each dot represents an individual F₂ mouse. NS, Not significant.

FIGURE 3

Interval map for granuloma size in S. mansoni-infected F₂ mice. The markers D4Mit203 (LOD of 3.4) and D17Mit82 (LOD of 6.0) were significantly linked to granuloma size. LOD scores are represented on the y-axis and chromosomal positions on the x-axis. Vertical dashed lines represent chromosomal breaks. Results are based on analysis performed by J/QTL (5000 permutations).

FIGURE 4

Segregation of granuloma size and cytokine production linked to peak markers D4Mit203 (A) and D17Mit82 (B) in F₂ mice. Average granuloma size was calculated for individual mice within the indicated genotypes before grouping. For the cytokines, each bar represents mean values ± SEM of mice of each genotype. Statistically significant differences between groups are as indicated in graphs. For D4Mit203: SS, n = 38; SB, n = 70; BB, n = 42; and for D17Mit82: SS, n = 36; SB, n = 71; BB, n = 41.

FIGURE 5

Segregation of granuloma size in F₂ mice homozygous for the SJL or BL/6 allele or both at D4Mit203 and D17Mit82. Each bar represents mean granuloma size of mice per group. At least 20 granulomas were measured per mouse. NS, Not significant.

FIGURE 6

Granuloma size and cytokine production in SJL.B6-D4Mit203 and SJL.B6-D17Mit82 congenic mice. Congenic mice were produced as described in Materials and Methods. SJL.B6-D4Mit203 and SJL.B6-D17Mit82 congenic mice, or mice congenic for both intervals, had significantly reduced granuloma size (A) and IL-17 production (B) compared with SJL and littermate controls; however, these loci did not affect IFN-γ production (C). Granulomas were counted from individual mice and at least 20 granulomas were counted per mouse. Each dot represents an individual mouse. For cytokine values, each dot represents the average of triplicate determinations of individual mice. Black horizontal lines represent the mean of the group. *, p < 0.05; **, p < 0.01; ***, p < 0.001.