High parasite burdens cause liver damage in mice following Plasmodium berghei ANKA infection independently of CD8(+) T cell-mediated immune pathology - PubMed (original) (raw)
High parasite burdens cause liver damage in mice following Plasmodium berghei ANKA infection independently of CD8(+) T cell-mediated immune pathology
Ashraful Haque et al. Infect Immun. 2011 May.
Abstract
Infection of C57BL/6 mice with Plasmodium berghei ANKA induces a fatal neurological disease commonly referred to as experimental cerebral malaria. The onset of neurological symptoms and mortality depend on pathogenic CD8(+) T cells and elevated parasite burdens in the brain. Here we provide clear evidence of liver damage in this model, which precedes and is independent of the onset of neurological symptoms. Large numbers of parasite-specific CD8(+) T cells accumulated in the liver following P. berghei ANKA infection. However, systemic depletion of these cells at various times during infection, while preventing neurological symptoms, failed to protect against liver damage or ameliorate it once established. In contrast, rapid, drug-mediated removal of parasites prevented hepatic injury if administered early and quickly resolved liver damage if administered after the onset of clinical symptoms. These data indicate that CD8(+) T cell-mediated immune pathology occurs in the brain but not the liver, while parasite-dependent pathology occurs in both organs during P. berghei ANKA infection. Therefore, we show that P. berghei ANKA infection of C57BL/6 mice is a multiorgan disease driven by the accumulation of parasites, which is also characterized by organ-specific CD8(+) T cell-mediated pathology.
Figures
Fig. 1.
Large numbers of antigen-specific CD8+ T cells accumulate in the liver during ECM. (A) C57BL/6 mice (n = 5) were infected with P. berghei ANKA-luc, and 6 days later, when mice displayed ECM symptoms, CD8+ T cells in the brain, spleen, and liver were enumerated in these and naïve mice by flow cytometry. Data are representative of three independent experiments. (B) C57BL/6 mice (n = 5) were adoptively transferred by i.v. injection with 10,000 CD8+ MACS-purified, congenically marked CD45.1+ OTI cells and 2 h later were infected with P. berghei ANKA-Ova or left uninfected. Six days later, when infected mice displayed ECM symptoms, CD8+ CD45.1+ OTI T cells in the brain, spleen, and liver were enumerated by flow cytometry. Data are representative of two independent experiments. *, P < 0.05 (Mann-Whitney test) relative to corresponding naïve group.
Fig. 2.
The majority of liver-recruited CD8+ T cells express granzyme B during P. berghei ANKA infection. C57BL/6 mice (n = 5 or 6), were infected with P. berghei ANKA-luc or left naïve. At 6 days p.i., when infected mice displayed clinical scores of 1 or 2, splenic, liver, and brain CD8+ T cells were assessed for GzmB expression by intracellular staining and flow cytometric analysis. Representative fluorescence-activated cell sorter (FACS) histograms are gated on CD8+ TCRβ+ cells, and numbers alongside each gate depict the percentage of GzmB+ cells. **, P < 0.01 (Mann-Whitney test) relative to naïve spleen group. These data are summarized in the adjacent graph and are representative of two independent experiments performed.
Fig. 3.
Liver damage occurs in ECM prior to the onset of neurological symptoms. C57BL/6 mice (n = 6 to 11) were infected with P. berghei ANKA-luc, and 4, 5, or 6 days later plasma samples from these and naïve mice were assessed for liver ALT and AST levels. **, P < 0.01; ***, P < 0.001 (Mann-Whitney test, relative to naïve mice). Data are representative of three independent experiments.
Fig. 4.
Histopathological assessment of liver damage during P. berghei ANKA (PbA) infection. C57BL/6 mice (n = 5 or 6) were infected with P. berghei ANKA-luc or left naïve. (A) At 5 or 6 days after infection, liver-resident CD8+ T cells were enumerated by flow cytometry. *, P < 0.05; **, P < 0.01 (Mann-Whitney test, relative to naïve group). (B) At 5 or 6 days after infection, paraffin-embedded tissues were sectioned and stained with hematoxylin and eosin (upper panels) or with picrosirius red (lower panels). Collagen is depicted by red staining and malaria pigment by punctuate black staining in the lower panels. Bars, 10 μm. These data are representative of two independent experiments performed.
Fig. 5.
Parasites, not CD8+ or CD4+ T cells, mediate liver damage following P. berghei ANKA infection. C57BL/6 mice (n = 5) were infected with P. berghei ANKA-luc, and 3 days later infected mice, as well as naïve controls, were treated with control rat IgG, anti-CD4 MAb, anti-CD8β MAb, or artesunate. (A and B) A further 3 days later, when control rat IgG-treated mice were displaying ECM symptoms, mice were assessed for whole-body parasite burden (A) and liver parasite burden (B) by bioluminescence imaging. (C) Plasma samples were also tested for liver enzyme ALT and AST levels. **, P < 0.01 (Mann-Whitney test) for comparison of each treatment group to the control rat IgG treated group. These data are representative of two independent experiments.
Fig. 6.
Therapeutic artesunate treatment, but not CD8+ T cell depletion, ameliorates established liver damage following P. berghei ANKA infection. C57BL/6 mice (n = 5) were infected with P. berghei ANKA-luc, and 6 days later infected mice displaying ECM symptoms, as well as naïve controls, were treated with control rat IgG, anti-CD8β MAb, or artesunate. (A and B) Mice were then assessed at 1 day and 4 days after treatment for whole-body parasite burden (A) and liver parasite burden (B) by bioluminescence imaging. (C) Plasma samples were also tested for liver enzyme ALT and AST levels: **, P < 0.01 (Mann-Whitney test) for comparison of artesunate group to anti-CD8β MAb group at 4 days posttreatment. These data are representative of two independent experiments.
Fig. 7.
Perforin-deficient mice exhibit liver damage during P. berghei ANKA infection. C57BL/6 perforin-deficient mice (n = 5 or 6) were infected with P. berghei ANKA-luc or left naïve. Six days later, when these mice displayed no clinical symptoms of disease, plasma samples were tested for liver enzyme ALT and AST levels. **, P < 0.01 (Mann-Whitney test) relative to naïve perforin-deficient mice.
References
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