14 research outputs found
<i>S.</i> Typhimurium-infected mice have tissue inflammation and thrombosis, increased hematopoiesis, and decreased splenic iron.
<p>(A) Mouse liver, 6 weeks post-infection; inflammation and necrosis (arrow). (B) Mouse spleen, 3 weeks post-infection; extramedullary hematopoiesis (EMH; arrow, megakaryocytes), histiocytic infiltration (I) throughout the red pulp, and thrombus (T). H&E stain (A, B). (C) Spleen, mock-infected (left) and infected mouse (right), 3 weeks post-infection; markedly decreased ferric iron staining in red pulp. (D) Spleen, mock-infected (left) and infected mouse (right), 6 weeks post-infection; markedly decreased splenic ferric iron in red pulp. Perl's Prussian Blue stain (C, D). (E) Hemophagocytic macrophage in mouse spleen 3 weeks post-infection that had 10-fold more macrophages and 43-fold more 6N+ macrophages than control mouse spleen. CD11b (red), DAPI (blue), TER119 (green). N  =  endogenous macrophage nucleus, E1  =  nucleated erythrocyte, E2  =  non-nucleated erythrocyte. Confocal fluorescent micrograph. (F) Representative histogram overlay of TER119 expression on DAPI+ splenocytes from a mock-infected (red) and infected mouse (blue) 3 weeks post-infection. Filled gray histogram corresponds to the isotype control. The infected mouse had 11.5-fold more TER119<sup>med</sup> pro-erythroblasts and 5.5-fold more TER119<sup>high</sup> erythroblasts than the mock-infected mouse. (G) Mean numbers of TER119<sup>med</sup> and TER119<sup>high</sup> splenocytes from three mock-infected (white bars) and four infected (gray bars) mice. Mean number of TER119<sup>med</sup> pro-erythroblasts per spleen increased 6.8-fold in infected mice, while the mean number of TER119<sup>high</sup> cells, corresponding to all nucleated erythroblasts subsequent to the pro-erythroblast stage <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0009441#pone.0009441-Socolovsky1" target="_blank">[40]</a>, increased 3.6-fold. (<i>P</i><0.05) Error bars = SD. Original magnifications 100× (A–B), 200× (C–D), and 1000× (E).</p
Hematology of <i>S.</i> Typhimurium-infected mice: acute, then chronic active inflammatory response; microcytic anemia, persistent microcytosis.
<p>Mice were orally gavaged with 9.1×10<sup>8</sup> CFU of <i>S</i>. Typhimurium (n = 8) or sterile PBS (n = 7). Complete blood counts were monitored over 16 weeks. X  =  <i>S</i>. Typhimurium-infected mice; circle  =  mock-infected control mice. Mean and standard deviation are shown. (A) neutrophils, (B) monocytes, (C) lymphocytes, (D) hematocrit (HCT), (E) mean cell volume (MCV). *<i>P</i><0.05 (Student's <i>t</i>-test).</p
Clinico-pathologic features of HLH in <i>S.</i> Typhimurium-infected mice.
<p>PI indicates post-oral infection with 2.0×10<sup>9</sup> CFU <i>Salmonella enterica</i> serotype Typhimurium; C indicates mock-infected control mice.</p>*<p><i>P</i><0.05, Student's <i>t</i>-Test.</p>†<p>Independent experiment, same bacterial dosing and range for splenic bacterial CFU results.</p>‡<p>Formal diagnostic criteria for HLH per the Histiocyte Society guidelines <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0009441#pone.0009441-Henter1" target="_blank">[5]</a>.</p>§<p>Consistent with a diagnosis of HLH, and <sup>Π</sup> strong supportive evidence for HLH <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0009441#pone.0009441-Henter1" target="_blank">[5]</a>.</p
Vacuolation of diverse cell types in homozygous <i>Cln3</i><sup>Δ</sup>
<p><sup><b><i>ex7/8</i></b></sup><b> mice.</b> (A) Representative images are shown of Wright-Giemsa stained peripheral blood smears from <i>Cln3<sup>+/+</sup></i> and <i>Cln3</i><b><sup>Δ</sup></b><sup><i>ex7/8/</i><b>Δ</b><i>ex7/8</i></sup> littermate mice (scale bar = 10 µm). Note the presence of vacuoles in the cytoplasm of the dark blue stained peripheral blood lymphocyte. (B) Representative images are shown of H&E-stained sections of epididymis from 19-week-old <i>Cln3<sup>+/+</sup></i> and <i>Cln3</i><b><sup>Δ</sup></b><sup><i>ex7/8/</i><b>Δ</b><i>ex7/8</i></sup> littermate male mice (scale bar = 50 µm). A representative image of a section of mutant (<i>Cln3</i><b><sup>Δ</sup></b><sup><i>ex7/8/</i><b>Δ</b><i>ex7/8</i></sup>) epididymis immunostained for vacuolar ATPase (V-ATPase, green) and aquaporin-9 (AQP9, red), which highlight the apical (luminal) membrane of clear/narrow cells or principal cells, respectively (scale bar = 25 µm). (C) Representative TEM images of <i>Cln3</i><b><sup>Δ</sup></b><sup><i>ex7/8/</i><b>Δ</b><i>ex7/8</i></sup> epididymis cross-sections are shown. Note both the giant vacuoles and the multiple smaller vacuoles filling the cytoplasm of the clear cells. Also note the relative absence of electron-dense material inside the vacuoles. Scale bars, left panel = 10 µm; right panel = 2 µm. (D) Representative images of subunit c immunostained <i>Cln3<sup>+/+</sup></i> and <i>Cln3</i><b><sup>Δ</sup></b><sup><i>ex7/8/</i><b>Δ</b><i>ex7/8</i></sup> epididymis sections are shown. Asterisks (*) mark some of the large vacuoles. Scale bars = 50 µm. Blood smears and epididymides from at least 10 mice per genotype were analyzed in total, and abnormal vacuolation was observed in all of the <i>Cln3</i><b><sup>Δ</sup></b><sup><i>ex7/8/</i><b>Δ</b><i>ex7/8</i></sup> mice and in none of the wild-type or <i>Cln3<sup>+/</sup></i><sup><b>Δ</b><i>ex7/8</i></sup> mice.</p
Metabolic abnormalities in <i>Cln3</i><sup>Δ</sup>
<p><sup><b><i>ex7/8</i></b></sup><b> mice.</b> (A) Graphs depicting female (left) and male (right) mean body weight data from wild-type (diamonds), heterozygous (squares), and homozygous (triangles) <i>Cln3</i><b><sup>Δ</sup></b><sup><i>ex7/8</i></sup> mice at ages between 11 and 20-weeks are shown (n = 5–10 mice per genotype/sex/age). No significant genotypic differences were observed. Error bars represent SEM. (B) Mean ± SEM rectal body temperatures are shown for male (black bars) and female (gray bars) wild-type (<i>Cln3<sup>+/+</sup></i>), heterozygous (<i>Cln3<sup>+/</sup></i><sup><b>Δ</b><i>ex7/8</i></sup>) and homozygous (<i>Cln3</i><b><sup>Δ</sup></b><sup><i>ex7/8/</i><b>Δ</b><i>ex7/8</i></sup>) littermate mice are shown. Rectal body temperatures, which were measured at rest, were slightly elevated in male and female, heterozygous and homozygous <i>Cln3</i><b><sup>Δ</sup></b><sup><i>ex7/8</i></sup> mice, compared to wild-type mice. *, p<0.001 (heterozygous versus wild-type, homozygous versus wild-type). (C) Mean ± SEM values for minimum oxygen consumption (ml/hr) are shown for male (black bars) and female (gray bars) wild-type (<i>Cln3<sup>+/+</sup></i>), heterozygous (<i>Cln3<sup>+/</sup></i><sup><b>Δ</b><i>ex7/8</i></sup>) and homozygous (<i>Cln3</i><b><sup>Δ</sup></b><sup><i>ex7/8/</i><b>Δ</b><i>ex7/8</i></sup>) littermate mice are shown. Minimum oxygen consumption was elevated in male and female heterozygous and homozygous <i>Cln3<sup>Δex7/8</sup></i> mice, compared to wild-type mice. 5–10 mice per group (genotype/sex) were analyzed. *, p<0.001 (heterozygous versus wild-type, homozygous versus wild-type).</p
Heart analysis of <i>Cln3</i><sup>Δ</sup>
<p><sup><b><i>ex7/8</i></b></sup><b> mice.</b> (A) The bar graph depicts normalized heart weights for wild-type (<i>Cln3<sup>+/+</sup></i>), heterozygous (<i>Cln3<sup>+/</sup></i><sup><b>Δ</b><i>ex7/8</i></sup>), and homozygous (<i>Cln3</i><b><sup>Δ</sup></b><sup><i>ex7/8/</i><b>Δ</b><i>ex7/8</i></sup>) littermate 19–20 week old mice. Normalized heart weights represent a ratio of heart weight (mg = milligrams)/body weight (g = grams). Normalized heart weights were slightly increased in heterozygous <i>Cln3</i><b><sup>Δ</sup></b><sup><i>ex7/8</i></sup> mice, and more so in homozygous <i>Cln3</i><b><sup>Δ</sup></b><sup><i>ex7/8</i></sup> mice, compared to wild-type littermates. ANOVA analysis suggested a significant genotype effect (p<0.05). (B) Representative micrographs of hematoxylin and eosin (H&E) stained heart sections from wild-type (<i>Cln3<sup>+/+</sup></i>, n = 8) and homozygous (<i>Cln3</i><b><sup>Δ</sup></b><sup><i>ex7/8/</i><b>Δ</b><i>ex7/8</i></sup>, n = 10) littermate 19–20 week old mice are shown, which do not obviously differ from one another in their morphology. Scale bar = 100 µm. (C) Representative micrographs are shown of α-subunit c immunostained heart sections from 19-week old <i>Cln3<sup>+/+</sup></i> and <i>Cln3</i><b><sup>Δ</sup></b><sup><i>ex7/8/</i><b>Δ</b><i>ex7/8</i></sup> littermate mice. Note the abundance of subunit c-immunopositive deposits in the <i>Cln3<sup>?ex7/8/</sup></i><sup><b>Δ</b><i>ex7/8</i></sup> section. Only sparse punctate subunit c immunostaining is present in the <i>Cln3<sup>+/+</sup></i> section. Scale bar = 200 µm. Inset scale bar = 25 µm.</p
T cell frequencies in peripheral blood from <i>Cln3</i><b><sup>Δ</sup></b><sup><i>ex7/8</i></sup> mice.
<p>The frequencies of T-cells [% T cells (CD45+)], the ratios of CD4+/CD8+ T cells, and the percentage of Ly6c+ cells among the CD8+ and CD4+ T cell populations, determined by flow cytometry, are shown for female and male wild-type (<i>Cln3<sup>+/+</sup></i>), heterozygous (<i>Cln3<sup>+/</sup></i><sup><b>Δ</b><i>ex7/8</i></sup>), and homozygous (<i>Cln3<sup>Δex7/8/</sup></i><sup><b>Δ</b><i>ex7/8</i></sup>) littermate mice. p values, determined in a two-tailed, unpaired Student’s t-test of the heterozygous <i>Cln3</i><b><sup>Δ</sup></b><sup><i>ex7/8</i></sup> values versus wild-type (<i>Cln3<sup>+/+</sup></i>) values, or homozygous <i>Cln3</i><b><sup>Δ</sup></b><sup><i>ex7/8</i></sup> values versus wild-type (<i>Cln3<sup>+/+</sup></i>) values, are shown. Bold typeface highlights parameters that were significantly different versus wild-type controls. Samples from 9–10 mice per group (genotype/sex) were analyzed, as indicated.</p
Subunit c immunohistochemistry of major hematopoietic tissues from 12-week-old <i>Cln3</i><sup>Δ</sup><sup><i>ex7/8</i></sup> mice.
<p>Representative images from bone marrow brush cytology, tibia cross-sections (‘Bone marrow histology’), and liver and spleen sections immunostained for subunit c are shown for wild-type (<i>Cln3<sup>+/+</sup></i>) and homozygous mutant (<i>Cln3</i><b><sup>Δ</sup></b><sup><i>ex7/8/</i><b>Δ</b><i>ex7/8</i></sup>) mice (n = 2−5 mice per tissue/genotype). Inset in <i>Cln3</i><b><sup>Δ</sup></b><sup><i>ex7/8/</i><b>Δ</b><i>ex7/8</i></sup> bone marrow cytology panel (top right panel) shows a sea-blue histiocyte (arrows) from an H&E stained preparation. Sea-blue histiocytes were not found in wild-type bone marrow cytology preparations. Brown stain reflects subunit c-positive storage material, which is most prominent in cells that appear morphologically consistent with macrophages. Arrowheads mark examples of subunit c filled Kupffer cells in liver, also a macrophage lineage cell. Insets in histology panels show lower power magnification of subunit c immunostain. Scale bars = 25 µm.</p
Summary of neurological and behavioral testing of <i>Cln3</i><b><sup>Δ</sup></b><sup><i>ex7/8</i></sup> mice.
<p>A summary of genotypic differences observed in the neurological and behavioral screens is shown, with male and female results shown separately. Ages at which the indicated tests were performed are also shown.</p
Bone marrow analysis of <i>Cln3</i><sup>Δ</sup>
<p><sup><b><i>ex7/8</i></b></sup><b> mice.</b> Representative images are shown of Wright-Giemsa-stained bone marrow brush cytology, H&E stained sections of formalin-fixed, paraffin embedded tibias, and iron stained brush cytology, from wild-type (<i>Cln3<sup>+/+</sup></i>) and homozygous mutant (<i>Cln3</i><b><sup>Δ</sup></b><sup><i>ex7/8/</i><b>Δ</b><i>ex7/8</i></sup>) mice (n = 3 mice per genotype). Stained iron appears blue. Note the reduced amount of stained iron in <i>Cln3</i><b><sup>Δ</sup></b><sup><i>ex7/8/</i><b>Δ</b><i>ex7/8</i></sup> marrow, compared to wild-type marrow. Arrow, erythroid element; arrowhead, myeloid element; asterisk, megakaryocyte. Scale bars, top and bottom panels = 25 µm; middle panels = 100 µm.</p