Oxidative damage of DETC is restricted to amastigotes in the phagosome.

<p>Transmission electron microscopy of human macrophages infected with <i>Leishmania amazonensis</i>. Human macrophages were infected with <i>Leishmania amazonensis</i> (5∶1 ratio) for 4 h and then treated with DETC (2 mM) for 10 h. Untreated macrophages displayed numerous well-preserved parasites (A and B). DETC-treated macrophages presented remarkably damaged parasites (C and D), whereas host cell and parasite mitochondria were not injured (white and black arrows, respectively). P indicates intracellular parasites and N indicates host cell nucleus, bars represent 2 µm (A), 1 µm (B and C) or 0.5 µm (D). Images are representative of 2 independent experiments.</p

Dose-dependent leishmanicidal effect of DETC in <i>Leishmania</i>-infected human macrophages.

<p>Human monocyte-derived macrophages were infected with <i>Leishmania amazonensis</i> promastigotes (5∶1 ratio). (A) After infection, cells were treated with increasing concentrations of DETC and the number of intracellular amastigotes (symbols) was quantified as described in Material and <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0014394#s2" target="_blank">Methods</a>. Each bar represents the mean ± SEM of 100 cells counted in duplicate (One-way ANOVA, ***<i>p</i> = 0.0001; post test for linear trend, ***<i>p</i><0.001) representative of two different donors. (B) and (C) <i>Leishmania amazonensis</i>-infected macrophages were treated 48 h after infection with 2 mM of DETC. (B) The number of intracellular amastigotes was quantified as described in Material and <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0014394#s2" target="_blank">Methods</a>. Results are expressed as number of amastigotes/100 cells. Each bar represents the mean ± SEM of 4 donors (Paired <i>t</i> test, *<i>p</i> = 0.046). (C) Intracellular survival of <i>Leishmania amazonensis</i> amastigotes was quantified by transformation of proliferating extracellular motile promastigotes in Schneider's medium. Each bar represents the mean ± SEM of 4 donors (Paired <i>t</i> test, ***<i>p</i> = 0.0002).</p

Leishmanicidal effect of DETC in murine macrophages <i>in vitro</i>.

<p>(A) Murine (BALB/c) macrophages were infected with <i>Leishmania braziliensis</i> (5∶1 ratio) and treated with DETC (2 mM). Number of intracellular amastigotes was quantified as described in Materials and <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0014394#s2" target="_blank">Methods</a>. Each bar represents the mean ± SEM of six (Mann Whitney test, **<i>p</i> = 0.0049). (B) Murine (BALB/c) macrophages were treated with DETC (2 mM) in the presence of hydroxylamine (0.5 mM) and supernatant was collected at 48 h. Superoxide production was quantified as described in Materials and <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0014394#s2" target="_blank">Methods</a>. Each bar represents the mean ± SEM of six (Mann Whitney test, **<i>p</i> = 0.0043).</p

Oxidative damage of DETC in axenic promastigotes is restricted to mitochondria.

<p>Transmission electron microscopy of promastigotes form of <i>Leishmania amazonensis</i> in axenic culture treated with DETC. Promastigotes were treated with DETC (0.2 mM) for 1 h. (A) Untreated promastigotes displayed a well-preserved cytoplasm and electron-dense mitochondria with normal cristae. (B) DETC-treated promastigotes presented a well-preserved cytoplasm, besides enlarged mitochondria with reduced electron density, often with parallel or circular cristae (thin arrow). Dense core-containing mitochondria reacted positively for calcium oxalate (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0014394#s2" target="_blank">Methods</a>) by cytochemical detection (large arrow). (C) The use of NAC partially reverted the effects of DETC upon parasite mitochondria (large arrows with white margins). Images are representative of 2 independent experiments.</p

Zika Virus Infection and Stillbirths: A Case of Hydrops Fetalis, Hydranencephaly and Fetal Demise - Fig 1

<p><b>Axial ultrasound views of the fetus at the 30^th gestational week showing (A) Cranium with severe microcephaly (215mm) and hydranencephaly; (B) Posterior fossa with destruction of the cerebellar vermis (wide arrow) and nuchal edema (thin arrow); (C) Thorax with bilateral pleural effusions (arrow); and (D) Abdomen with ascites (wide arrow) and subcutaneous edema (thin arrow).</b></p

AA treatment dose-dependently induces cell death in HTLV-1-infected cell lines.

<p>HTLV-1-infected CD4+ T-cell lines were cultured for 48 hours with no treatment (NT), low-, intermediate-, high-dose AA (10, 50, 100 µg/ml) or IFN-α (1000 IU/ml). Flow cytometric quantification of cells with DNA degradation (Hoechst 33342-positive subdiploid cells) and proliferating cell nuclear antigen (PCNA)-positive cells are shown in the <i>y</i>-axis for both (A) MT-4 (n = 4) as well as (B) MT-2 cells (n = 3), whereas the treatment conditions are shown in the <i>x</i>-axis. ANOVA with Bonferroni post-test for multiple testing was used and the p-values are indicated by asterisks (^*<0.05, ^**<0.01 and ^***<0.001). (C) Confocal microscopy images of Hoechst 33342-stained MT-2 cells are shown for untreated and high-dose AA-treated MT-2 cells. Nuclear condensation (c) was observed for the AA-treated cells, whereas normal nuclei and mitosis (m) were observed for the untreated cells.</p

High-dose AA, but not IFN-α, exerts strong antiproliferative effects in HAM/TSP PBMCs.

<p>(A) Lymphoproliferation of normal donors (NDs) PBMCs (n = 4) and HAM/TSP PBMCs (n = 3) was quantified by [³H]thymidine incorporation (cpm), performed in triplicate. NDs and HAM/TSP PBMCs were cultured for 5 days with no treatment (NT), high-dose AA (100 µg/ml) or IFN-α (1000 IU/ml). Thymidine incorporation is shown in the <i>y</i>-axis, whereas the treatment conditions are shown in the <i>x</i>-axis. ANOVA with Bonferroni post-test for multiple testing was used and the p-values are indicated by asterisks (^*<0.05, ^**<0.01 and ^***<0.001). (B) NDs PBMCs (n = 3) and HAM/TSP PBMCs (n = 6) were cultured in the absence or presence of low-dose AA (10 µg/ml), high-dose AA (100 µg/ml) or IFN-α2A (1000 IU/ml) for 72 hours. Direct cell counts of viable cells were quantified by trypan blue dye exclusion. ANOVA with post-test for linear trend was used and the p-values are indicated by asterisks (^*<0.05, ^**<0.01 and ^***<0.001).</p

Overview of the significant down-regulated genes and the top 20 of the most significant up-regulated genes by IFN-α in MT-2 cells.

<p>Overview of the significant down-regulated genes and the top 20 of the most significant up-regulated genes by IFN-α in MT-2 cells.</p

AA, but not IFN-α, induces cell death in HAM/TSP PBMCs.

<p>HAM/TSP PBMCs (n = 6) were treated for 72 hours in the absence or presence of low-dose AA (10 µg/ml), high-dose AA (100 µg/ml) or IFN-α2A (1000 IU/ml). (A) The percentage of cell death was quantified by trypan blue dye exclusion and is shown in the <i>y</i>-axis, whereas the treatment conditions are shown in the <i>x</i>-axis. The one-sample t-test p-values are indicated by asterisks (^*<0.05, ^**<0.01 and ^***<0.001). (B) Fluorescence microscopy images of Hoechst 33342-stained PBMCs are shown for untreated (NT) and high-dose AA-treated PBMCs of one representative HAM/TSP patient (n = 3). Nuclear condensation and DNA fragmentation (f) was observed for AA-treated PBMCs.</p

Overview of the top 20 of the most significant down- and up-regulated genes by high-dose AA (100 µg/ml) in MT-2 cells.

<p>Overview of the top 20 of the most significant down- and up-regulated genes by high-dose AA (100 µg/ml) in MT-2 cells.</p