Fig 1.
E. multilocularis infection induces progressive liver pathological damage in wild-type mice.
(A) Comparison of absolute liver weight between infected wild-type (WT) mice and uninfected control (CON) group at different time points post-infection (2, 12, 24 weeks). (B) Liver-to-body weight ratio (liver weight/body weight × 100%). (C) Serum levels of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) activity. Data are presented as mean ± SD, n = 5 mice/group. *P < 0.05, **P < 0.01, ***P < 0.001, ns, P > 0.05. Uninfected control group vs infected WT group.
Fig 2.
E. multilocularis infection activates the hepatic CCR8/CCL1 axis and induces FOXP3 expression in the late stage.
(A) Representative immunohistochemical images showing protein expression of CCR8, CCL1, and FOXP3 in liver tissue at 24 weeks post-infection (400x magnification). (B) Quantitative analysis of the positive staining area (percentage). (C) qPCR analysis of the relative mRNA expression levels of CCR8, CCL1, and FOXP3 in the liver at different time points post-infection (normalized to GAPDH). (D) Immunofluorescence double staining shows co-localization of FOXP3 (green) and CCR8 (red) in lymphocytes surrounding liver lesions in the late stage of infection; DAPI (blue) stains nuclei, with an enlarged view. Data are presented as mean ± SD, n = 5 mice/group, *P < 0.05, **P < 0.01, ***P < 0.001, ns, P > 0.05. Uninfected control group vs infected WT group.
Fig 3.
CCR8+ T cell subsets expand and exhibit a functionally suppressed phenotype in the liver after E. multilocularis infection. (A) Flow cytometric analysis showing the proportions of CD4+ CD25+ FOXP3+ Tregs and their CCR8 + subset in the liver at different time points post-infection. (B) Proportions of CCR8+ cells among total CD3+, CD4+, and CD8+ T cells. (C) Comparison of CD4+ and CD8+ subset distribution between CCR8+ and CCR8- T cells. (D) Comparison of Granzyme B and Perforin expression between CCR8+ and CCR8- CD4+ and CD8+ T cells. Data are presented as mean ± SD, n = 5 mice/group. *P < 0.05, **P < 0.01, ***P < 0.001, ns, P > 0.05. (A)-(B) is compared between the uninfected control group and infected WT groups.
Fig 4.
CCR8+ T cells exhibit impaired effector function and a regulatory polarization profile. (A) Comparison of perforin, IFN-γ, TNF-α, and IL-2 expression between CCR8+ and CCR8- CD4+ and CD8+ T cells. (B) Differences in the expression of Th1/Th2/Th17 and related regulatory cytokines (IL-4, IL-17A, IL-10, TGF-β1) between the two cell populations. Data are presented as mean ± SD, n = 5 mice/group. *P < 0.05, **P < 0.01, ***P < 0.001, ns, P > 0.05.
Fig 5.
CCR8 deficiency alleviates E. multilocularis-induced liver pathological damage and downregulates immunosuppressive molecule expression.
(A) Representative in vivo liver ultrasound images of infected wild-type (WT) and CCR8 knockout (KO) mice at 12 weeks post-infection. Black arrows indicate parasitic lesions. (B) Quantitative analysis of lesion volume based on ultrasound images (n = 5 mice/group). (C) Absolute liver weight (left) and liver-to-body weight ratio (right, liver weight/body weight × 100%) at 12 and 24 weeks post-infection. The significant reduction in liver weight and liver-to-body weight ratio indirectly reflects the reduced burden of space-occupying lesions in infected CCR8-KO mice due to controlled parasite infiltration and growth. (D) Serum ALT levels at different time points post-infection. (E-F) Representative immunohistochemical images and quantitative analysis showing protein expression of CCL1 and FOXP3 in the livers of infected WT and KO mice at 24 weeks post-infection. (G) qRT-PCR analysis showing the relative mRNA expression levels of CCL1 and FOXP3 in the livers of infected WT and KO mice at 24 weeks post-infection (normalized to GAPDH). All data are presented as mean ± SD. n = 5 mice/group. *P < 0.05, **P < 0.01, ***P < 0.001, ns, P > 0.05, infected WT group vs. infected KO group.
Fig 6.
CCR8 deficiency enhances the effector functions of liver T cells.
(A) Flow cytometric analysis showing the proportions of liver CD4+ and CD8+ T cells from infected WT and KO mice secreting IFN-γ and TNF-α at different time points post-infection. (B) Flow cytometric analysis showing the proportions of liver CD4+ and CD8+ T cells from infected WT and KO mice secreting Granzyme B and Perforin at different time points post-infection. (C) Proportions of CD4+ and CD8+ T cells secreting IL-2 from infected WT and KO mice. Data are presented as mean ± SD, n = 5 mice/group. *P < 0.05, **P < 0.01, ***P < 0.001, ns, P > 0.05. Infected WT group vs infected KO group.
Fig 7.
Effect of CCR8 deficiency on Th2/Th17 and regulatory cytokine expression.
(A) Flow cytometric analysis showing the proportions of CD4+ T cells and CD8+ T cells secreting IL-4 and IL-17A. (B) Proportions of CD4+ T cells and CD8+ T cells secreting IL-10 and TGF-β1. Data are presented as mean ± SD, n = 5 mice/group. *P < 0.05, **P < 0.01, ***P < 0.001, ns, P > 0.05. Infected WT group vs infected KO group.