Fig 1.
Anemia caused by L. donovani infection.
Mice were infected with 1x107 L. donovani promastigotes. After 12 and 24 weeks, infected mice as well as age-matched naïve mice were sacrificed to examine organ weights and parasite burden of the spleen (A and B) and liver (C and D), hematocrit (E), hemoglobin (F), peripheral blood red blood cell counts (G), serum erythropoietin levels (H), and proportion of polychromatic erythrocytes in peripheral blood (J). The open symbols or bars represent naïve mice and closed symbols or bars represent infected mice. The mean and SD of at least 3 mice in each group are shown. These are representative of three independent experiments with similar results. P values by two-way ANOVA followed by Bonferroni’s multiple comparison test (for E) or unpaired t test (for F, G, H and J) are shown. ns, not significant. Fig 1I shows representative microscopic images of Giemsa-stained smears of peripheral blood from infected mice. Arrows indicate polychromatic erythrocytes. Scale bar, 20 μm.
Fig 2.
Hemophagocytosis in the spleen of L. donovani-infected mice.
(A, B) Low magnification images of HE-stained sections of the red pulp of spleens from naïve (A) or L. donovani-infected mice (B). Hemophagocytes engulfing multiple erythrocytes (arrows) were found in the spleen of the infected mice, whereas such cells were not found in naïve mice. (C) A high magnification image of an HE-stained section of the spleen from infected mice. L. donovani amastigotes (arrowheads) were often found in hemophagocytes. Scale bars, 20 μm.
Fig 3.
Heavy infection of hemophagocytes by Leishmania parasites.
The number of amastigotes per macrophage/hemophagocyte in the spleen of L. donovani-infected mice was analyzed by microscopy of HE-stained tissue sections. Host cells were categorized, based on the number of parasites per cell, into the groups none (0 amastigotes), low (1–10 amastigotes), moderate (11–20 amastigotes) and high (more than 20 amastigotes). The graph shows percentages of cells with no, low, moderate and high infection in either total macrophages or hemophagocytes only.
Fig 4.
The multinuclear character of hemophagocytes in the spleen of L. donovani-infected mice.
(A) A representative light microscopic image of MGCs engulfing erythrocytes in a thin section of a spleen from L. donovani-infected mice (24 weeks p.i.). Scale bar, 20 μm. (B) Proportion of mononuclear and multinuclear cells in the total macrophage and hemophagocyte only populations.
Fig 5.
Characterization of heavily infected and multinucleated macrophages by immunohistochemistry.
Sections of the spleen of L. donovani-infected mice at 24 weeks post-infection were stained with anti-F4/80 (A), MOMA-2 (B) or anti-CD11b (C) antibody, followed by counterstain with hematoxylin. Cells in the red pulp with positive signal are shown. Parasites are visualized as blue dots. Scale bars, 20 μm.