Skip to main content
. 2017 Dec 7;15:97–108. doi: 10.1016/j.redox.2017.11.024

Fig. 3.

Fig. 3

HIV-1 Tat protein induces mitochondrial OS in B-cells. (A) Tat-induced ROS production is diminished by the mitochondrial ROS production inhibition. B-cells were treated with 250 ng/ml Tat alone or with 10 µM mitochondrial complex I inhibitor rotenone or 20 ng/ml mitochondria-targeting antioxidant SkQ1 or 10 µM uncoupler of oxidative phosphorylation DNP for 6 h and stained with DHE. The Y axis shows ROS production calculated from a minimum of 100 cells examined for each condition and compared to the negative control for which the obtained basal value was set as 1. The experiment was carried out on blood from three different healthy donors. (B) Representative images of MitoSOXRed-stained B-cells. Hoechst is shown in blue (nuclear staining), MitoSOX in red. Scale Bar = 10 µm. (C) Tat induces superoxide anion production in mitochondria of B-cells. B lymphocytes treated with 250 ng/ml Tat alone or with 20 ng/ml SkQ1 were stained with MitoSOXRed and visualized by fluorescence microscopy 6 h after treatment. ROS production was quantified as described in Materials and Methods. The Y axis represents the proportion of cells with MitoSOXRed staining in Tat-treated B-cells compared to the untreated control, for which the obtained basal value was set as 1. The values were calculated from a minimum of 100 cells examined for each condition. The experiment was carried out on blood from two different healthy donors. (D) SkQ1 inhibits Tat-induced DD. B-cells were treated with 250 ng/ml Tat alone or together with 20 ng/ml SkQ1 and then fixed at 6 h post-treatment and immunostained for γH2AX. The Y axis shows the proportion of γH2AX-positive Tat-treated B-cells compared to the untreated controls, for which the obtained basal value was set as 1. The number of γH2AX positive cells was calculated from a minimum of 300 cells examined per experimental point. The experiment was carried out on blood from two healthy donors. All data are expressed as the mean±SEM. The statistical analyses were carried out by the one-way ANOVA test. The statistical significance was calculated vs. untreated controls; ***: p value<0.001.